Microbiology Revision E6.5

Microbiology Revision 1 01 1
MICROBIOLOGY REVISION 1 ----- Active space -----
Stains 00:00:57
Stains Used for

Differential stain.
Gram stain Divides bacteria into gram positive (GP) &
gram negative (GN).
Acid fast stain Differential stain for mycobacteria.
Cold version of Ziehl-Neelsen stain.
Kinyoun stain
concentration of carbolic acid in 10 stain.
Acid fast stain for demonstrating
Fite Faraco stain
M. leprae.
Negative stain (India ink/Nigrosin) Capsules, spirochetes.
Albert, Neisser, Ponder stain volutin granules of Corynebacterium.
Ashby stain Spores of Bacillus, Clostridium etc.
Silver stains :
Levaditi (For tissue section) &
Spirochetes.
Fontana (For fluid specimen)
Warthin starry silver stain (Black) H. pylori, Bartonella.
ZN stain showing beaded, Fite Faraco stain showing Albert stain : Purplish blue
acid fast M. tuberculosis M. leprae in bunches. metachromatic granules of
(reddish pink rods) C. diphtheria.
Silver impregnation stain Warthin starry stain Negative stain showing

showing spirals of T.pallidum. showing H.Pylori. Cryptococcus.
Microbiology Revision • v1.0 • Marrow 6.5 • 2023

2 01 Microbiology
----- Active space ----- Steps of gram stain :

Crystal violet (Primary stain)
Stains cytoplasm purple/blue
lodine (Mordant)
Cells remain purple/blue
Alcohol/acetone (Decolorizer)
GP : Remain purple/blue GP cocci
GN : Colorless
Safranin (Counter stain) GN rods
GP : Remain purple/blue
Gram stain
GN : Pink/red
Steps of acid-fast stain :

Reagents of gram stain cannot stain the cytoplasm of acid-fast bacilli (AFB)
d/t lipid layers.
Carbol fuchsin (Primary stain)
All bacteria take up primary stain
Intermittent heating to allow the stain penetrate the lipid rich wall
20% sulphuric acid or 3% alchohol (Decolorizer)

AFB : Retain primary stain
Non acid-fast : Take up counter stain
Methylene blue/malachite green (Counter stain)
AFB : Retain primary stain.

Non acid-fast : Take up counter stain.
Acid fast staining :
Post methylene blue

Acid fast vs partially acid fast structures : ----- Active space -----
Acid-fast structures Partially acid-fast structures

Retain 1 stain with 20% H2SO4 :
0
Retain 10 stain with :
1. M. tuberculosis : Retain 10 stain with 3 % 1. 5 % H2SO4 :
acid alcohol (Alcohol fast). • M. leprae.
2. Atypical mycobacteria : • Oocysts of Cyclospora,
Do not retain 1°stain with 3 % acid Cryptosporidium, Cystoisospora.
alcohol (Non-alcohol fast). 2. 0.5 - 1 % H2SO4:
• Nocardia, Legionella micdacei .
3. 0.25 - 0.5 % H2SO4 :
• Bacterial spores, head of sperm.
Anatomy of Bacteria 00:10:13
Cell wall :
Differences between cell wall of GP & GN bacteria :
Features GP GN
1. Peptidoglycan :
Thick (50-100
Layers of murine monomers : Thin (2 layers).
layers).
• Made of alternate molecules of N-acetyl glu- Can be
Cannot be
cosamine & N-acetyl muramic acid. decolorized.
decolorized.
• Cross-linked by peptides.
Aromatic & sulfur containing amino acids (AA) in pep-
Absent Present
tide cross-linkage
2. Techoic acid Present Absent
3. Outer membrane Absent Present
4. Porins Absent Present
5. LPS (Lipopolysaccharide)/endotoxin Absent (Except in
Present
Listeria).
6. Periplasmic space Absent Present
LPS
Outer membrane
Peptidoglycan
Peptidoglycan
periplasmic space
Cytoplasmic
Cytoplasmic
membrane
membrane
Cell wall : GP Cell wall : GN

4 01 Microbiology
----- Active space -----

Capsule :
Polysaccharide in all except polypeptide capsule in B. anthracis.
Quellung/Neufeld reaction : Capsular swelling on adding anti-capsular
anti-serum.
Capsulated bacteria (Some Bacteria Have Killer And Mean Capsules) :
• S. pneumonia. • B. Anthracis.
• B .fragilis. • Meningococcus.
• H. influenzae. • Clostridium perfringens.
• Klebsiella.
Slime layer :
Help in biofilm formation → Prevent the entry of antibiotics (mechanical
barrier).
Endotoxin/LPS :
• Released only on cell lysis.
• Heat stable.
• Low antigenicity.
• 3 parts : Lipid A (Effects), core polysaccharide,
& somatic O-antigen.
• Binding of lipid A to TLR-4 (Toll-like receptor) →
Release of IL-1, 6, TNF α.
Parts of LPS
• Constant symptoms of endotoxemia
(Fever, hypotension, DIC, intravascular permeability) (D/t lipid A).
Exotoxins :
• Heat labile proteins.
• Secreted by both GP & GN.
• Toxoidable.
• Can be encoded on chromosome/plasmids/bacteriophages.
• Phage-mediated toxins formed by lysogenic conversion :
• Shiga like toxin of enterohemorrhagic E.coli (Verotoxin).
• Botulinum toxin C & D.
• Cholera toxin.
• Diphtheria toxin.
• Erythrogenic toxin A & C of Streptococcus pyogenes (Super antigens).

Modes of actions of toxins : ----- Active space -----
MOA Examples
• Cholera toxin.
• Anthrax toxin.
cAMP
• Enterotoxigenic E.coli (ETEC) labile toxin.
• Pertussis toxin & adenyl cyclase toxin of B. pertussis.
• ETEC stable toxin.
cGMP
• EAST-1 (Entero aggregative stable toxin).
By ADP ribosylation of EF-2 (Elongation factor) :
• Diphtheria toxin.
Inhibiting
• Exotoxin A of Pseudomonas aeruginosa.
protein
By cleavage of 60S ribosome :
synthesis
• Shiga toxin.
• Shiga-like toxin.
• Botulinum toxin → Inhibits AcH release.
• Tetanospasmin → Inhibits GABA & glycine release.
Neurotoxins
• S. aureus enterotoxin & B. cereus emetic toxin (Cereulide) →
Vagus nerve stimulation & vomiting centre.
Flagellum :
Monotrichous : Vibrio, P. aeruginosa
• Protein (Highly antigenic).
• Types of flagella :
Lophotrichous : H. pylori
Amphitrichous : Campylobacter, Spirillum
Peritrichous : Enterobacteriaceae, Clostridium, Listeria etc
Types of motility :
• Darting motility : Vibrio, Campylobacter.
• Corkscrew motility : Treponema pallidum (Endo flagella).
• Stately motility : Salmonella.
• Tumbling (End-on-end motility) : Listeria.
• Differential motility (motile at room temperature : 250C & non-motile at 370C)
: Listeria, Yersinia enterocolitica & Y. pseudotuberculosis.

6 01 Microbiology

Swarming :
Seen in GP Seen in GN
• Proteus.
• C. tetani.
• V. alginolyticus.
• B. cereus
• V. parahaemolyticus.
Swarming
Fimbriae/Pili : Made of Pilin protein.
2 types :
Common pili Sex pili
Seen in GN. Seen in both GP & GN.
Help in adhesion. F (Fertility/sex) plasmid + Helps in conjugation
(Connection between F+ & F- bacteria). Fimbriae
Endospores :
• Dipicolinic acid (Core) → Provides heat resistance.
• keratin-like protein (Spore coat) → Resistance to chemical disinfectants.
Pathogenic spores
Bacillus Clostridium
• Non bulging. • Bulging.

• Forms in soil & culture. • Forms in soil, culture & human body.
Cultivation of Bacteria 00:30:54
Types of culture medium :

1. Enriched medium : Basal medium (for fastidious bacteria) + egg/blood/serum.
Blood : Serum :
• Blood agar. • Loeffler’s serum slope
• Chocolate agar. • PPLO medium
• Skirrow’s medium. Egg : LJ/Dorset egg medium → M. tuberculosis.
• Butzler medium.
• Thayer Martin medium.
2. Enrichment medium (liquid media) : Basal/enriched medium + Any selecting
agent (To inhibit unwanted bacteria) :
• Alkaline peptone water → Vibrios.
• Selenite F broth → Shigella, Salmonella.
• Tetrathionate broth → Salmonella.

3. Selective medium (solid medium) : Basal/enriched medium + Any selecting ----- Active space -----
agent (To inhibit unwanted bacteria) :

Selective media Selective for
• Macconkey agar
GN bacteria
• EMB (Eosin methylene blue agar)
• DCA (Deoxycholate citrate agar)
Salmonella
• XLD (Xylose lysine deoxycholate agar)
Shigella
• Hektoen enteric agar
• Wilson & Blair medium Salmonella
• Skirrow’s medium Campylobacter
• Butzler medium Helicobacter (Skirrow’s)
• TCBS agar (Thiosulfate citrate bile salts sucrose) : pH
Vibrio
8.2 - 8.4
• Modified Thayer Martin medium Meningococci
• New York city medium Gonococci
• BCYEA (Buffered charcoal yeast extract agar) Legionella
• Salt agar Staphylococcus
• PLET medium (Polymyxin lysozyme EDTA
B. anthracis
thallous acetate)
• MYPA (Mannitol egg yolk polymyxin agar) B. cereus
• Macleod medium
• Tinsdale medium Corynebacterium (Potassium
• Hoyles medium tellurite agar)
• Downes medium
• CIN (cefsulodin irgasan novobiocin) agar Yersinia
4. Differential medium : Basal/enriched medium + Differentiating agent.

Differential medium Differentiate
α/β/γ hemolytic bacteria :
α : Incomplete hemolysis (Green).
Blood agar
β : Complete hemolysis (Yellow).
γ : No hemolysis (No color).
MacConkey agar
Lactose & non-lactose fermenting bacteria.
EMB agar
E.Coli O157 : H7 → Pale colonies.
Sorbitol MacConkey medium All other E.Coli → Pink colonies.
Mannitol (S. aureus) & non-mannitol
Mannitol salt agar
fermenting staphylococcus.
Sucrose (V. cholera) & non-sucrose
TCBS fermenting (V. parahaemolyticus) Vibrios.

8 01 Microbiology
MacConkey agar :
α/ β/ γ hemolytic bacteria LF (pink) NLF
EMB agar : Purple colonies of S.aureus (lt) : Yellow colonies V.cholera (left) &
E.Coli with green metallic sheen Rest (rt) : Pale colored colonies V. parahemolytics (right)
of CONS
Transport medium :
Transport medium Bacteria
• Cary Blair (Universtal stool transport medium).
• VR (Venkatraman Ramakrishnan) medium. Vibrio
• Autoclaved seawater.
Stuart & Amie’s media Neisseria
Thioglycolate Anaerobes
Pike’s medium Streptococcus pyogenes
Classification of Bacteria 00:42:22
Based on gas requirements :

• Obligate aerobes : Brucella, Bordetella, M. tuberculosis, Nocardia,
P. aeruginosa.
• Obligate anaerobes (Catalase, peroxidase & superoxide dismutase absent) :
Lactobacillus, Clostridium, Bacteroides, Actinomycetes, Prevotella.
• Facultative anaerobes : Enterobacteriaceae, Vibrio, Hemophilus.
• Microaerophilic : Campylobacter, Helicobacter.
• Capnophilic : Pneumococci, H. influenzae, Neisseria, Brucella abortus.

Biochemical Reactions 00:46:00 ----- Active space -----
Tests :
Tests Analysis Image
Negative in (StASh) :
• Streptococcus.
Catalase test • Anerobes.
Positive test : Bubbles on
• Shigella dysenteriae type I.
adding H2O2
Negative in : CESS
Oxidase test
• Corynebacterium
(Cytochrome
• Enterobacteriaceae
oxidase C
• Staphylococcus
detection) Positive test : Purple
• Streptococcus
• Oxidative utilization (Aerobic
condition).
• Fermentative utilization
(both aerobic & anaerobic
Sugar utilization
condition). Hugh & Leifson’s oxidative
Hugh & Leifson’s oxidative (fermentative medium) -
fermentative medium is used Positive test : Yellow color
to differentiate them.
Positive in (PUNCH MSKB) :
• Proteus
• Ureoplasma
• Nocardia
• Cryptococcus
Urease test • H. pylori (Maximum urease
producer)
• Morganella Positive test : Pink color
• S. aureus & S. saprophyticus
• Klebsiella
• Brucella

10 01 Microbiology
----- Active space ----- Antibiotic Sensitivity Tests 00:50:03
Manual methods
1. Dilution method : 2. Disc diffusion method : 3. Epsilometer/E-test :

• Gold standard. • M/C used. • Combination of
• Serial concentrations • 2 methods : Kirby Bauer dilution & disc
of antibiotic added in (M/C) & Stokes methods. diffusion method.
the broth/agar & MIC • E-strip impregnated
determined. with graded
concentration of
antibiotic.
All are quantitative except disc diffusion (Qualitative).
Standardisations in antibiotic sensitivity :

• Cation-adjusted Muller Hinton agar/broth should be used.
• Standard inoculum of test bacterium used to raise the turbidity to
0.5 McFarland.
• Incubation temperature: 370C.
• Time of interpretation of results : 16-18 hours of incubation.
Zone of
inhibition
MIC : Epsilometer
Minimum inhibitory
concentration (MIC)
Kirby Bauer disc diffusion test
Test strain Control strain
Stokes method :
Zones of inhibition
compared b/w test &
control to
determine whether
it is resistant or
sensitive.

Intrinsic Resistance of Bacteria 00:54:41 ----- Active space -----
Results from genetic, structural or physiological state.

Helps in identification of certain bacteria in laboratories.
Organism : Intrinsic resistance to Mechanism
Pseudomonas : Sulfonamides,
trimethoprim, tetracycline, Lack of uptake
chloramphenicol.
Produce β lactamases that
Stenotrophomonas : Carbapenems destroy drug before it reaches
targets
Burkholderia cepacia : Polymyxin B Altered LPS structures
Lack PBP’s (Penicillin binding protein) to
Enterococci : Cephalosporins
which cephalosporin binds.
Enterococci : Aminoglycosides (AG) Lack of sufficient oxidative
Anaerobic : AG metabolism to drive uptake of AG.
Inability to anaerobically reduce drug
Aerobic : Metronidazole
to its active form.
Gram (+ve) : Aztreonam Lack PBP to which aztreonam binds.
Gram (-ve) : Vancomycin Lack of uptake via outer membrane.
Bacterial Growth Curve 00:58:05
Phase Description
• Adaptation phase, variable duration.
• Total count (TC) & viable count (VC) constant.
1. Lag phase • Bacteria : Metabolically active.
• Maximum size at the end of the phase.
• Exponential phase.
• TC & VC increase.
• Metabolically most active.
2. Log phase
• Maximum effect of antibiotic.
• Generation time (Population doubling time)can be
calculated.

12 01 Microbiology
----- Active space ----- • Replicating bacteria = Dying bacteria.

3. Stationary
• TC increase, VC constant.
phase
• Spores, exotoxins formed & antibiotics produced.
• Nutrients depleted.
• TC constant & VC decrease.
4. Death phase • Toxic metabolites ++
• Involution forms seen.
Bacterial growth curve
Bacterial Gene Transfer 01:01:12
Transformation Transduction Conjugation

• Uptake of • Transfer of • Bacteria with F plasmid →
soluble DNA genes via Encodes conjugation tube/sex
directly via cell bacteriophage. pilus.
wall. • M/C • Copy of F+ plasmid transferred via
• Bacteria : mechanism of the tube to F- bacteria.
Competent gene transfer. • M/C mechanism of spread of
bacteria. antibiotic resistance.
Bacteriophage F+ F-
F plasmid

Catalase negative bacteria : Catalase Test

“StASh”
• Streptococcaceae members.
• All Anaerobes.
• Shigella dysenteriae type 1.
H202 → H20+ O2 (Bubbles)
Oxidase negative bacteria : Oxidase test (Detection of

“CESS” cytochrome C oxidase)
• Corynebacterium.
• Enterobacteriaceae.
• Staphylococci.
• Streptococcaceae.
Gram positive bacteria
G+ve cocci G+ve bacilli
Staphylococcus Streptococcus Gram Not gram

• Catalase +ve. • Catalase -ve. stained stained
• Grape like clusters. • Chains/pairs.
Mycobacterium
Gram Positive Cocci 00:02:08
Staphylococcaceae :
• Catalase +ve; Oxidase -ve.
• Salt tolerant (7-10% salt).
• Non fastidious.

14 02 Microbiology

Coagulase test :
Slide coagulase test : Tube coagulase test :

Detects bound coagulase. Detects free coagulase.
Staphylococcaceae types :
Coagulase Negative
S. aureus
staphylococcus (CONS)
Slide & tube coagulase
Both positive Any one/both negative
test
Pigment Golden pigment -
Hemolysis β hemolytic Non hemolytic
Non mannitol fermenter
Mannitol fermentation Mannitol fermenter (MF)
(NMF)
β hemolytic, big colonies Golden yellow pigment of S. Yellow colonies of S. aureus

of S. aureus aureus (MF) & pale colonies of CONS
(NMF) in mannitol salt agar.
CONS
S. Saprophyticus S. epidermidis
• Resistance to Novobiocin. • Sensitive to Novobiocin.
• UTI in females with recent • Slime producing bacteria.
sexual activity. • Present as skin flora.
• M/C cause of honeymoon • Bacteremia in IV catheterized,
cystitis. surgical site infections (SSI),
prosthetic valve endocarditis.

Staphylococcus aureus 00:06:27 ----- Active space -----
Slide & tube coagulase +ve.

DNAse +ve, Phosphatase +ve.
Mannitol fermentation +ve.
Nutrient agar : Golden yellow pigment.
Sensitive to Novobiocin.
Blood agar : β hemolytic colonies.
Selective media : Salt agar, Ludlam medium.
Differential medium : Mannitol salt agar.
Virulence factors :
• Bound & free coagulase.
• Protein A : Used in coagglutination test to detect soluble antigens in clinical
specimens.
Toxins :
α hemolysin Paradoxical reactivation b/w 80-1000C.
β hemolysin Hot and cold phenomenon.
γ hemolysin Synergohymenotropic toxins (Bi-component membrane
PVL damaging toxins)
Panton Valentine Leucocidin (PVL) : Dermo necrotic & leucocidal.
S. aureus is the m/c cause of
• Folliculitis, stye, boil, furuncle, impetigo, bullous impetigo, ecthyma, cellulitis,
SSI.
• Osteomyelitis.
• Septic arthritis
• Hospital acquired pneumonia.
• Ventilator associated pneumonia.
• Infective endocarditis in IV drug users.
• Acute Infective endocarditis.
• Native valve infective endocarditis (NVE).
• Early & late prosthetic valve endocarditis (as a single species).
Note :
Most common cause of (As a group)
1. Early Prosthetic valve endocarditis : CoNS.
2. Late Prosthetic valve endocarditis : Viridans group of streptococci.

16 02 Microbiology

Toxin mediated syndromes :
1. Enterotoxins :
Heat stable toxins.
Causes staphylococcal food poisoning (Short incubation period : 6 to 8 hours).
Risk factor : H/o ingestion of meat/milk products/pastries/potato salads/
coleslaw.
2. Exfoliative (Epidermolytic) toxins :

EF A, B causes Staphylococcal scalded skin syndrome (SSSS)/(Ritter’s
sydrome) or bullous impetigo.
3. TSST-1/SE-F :
Superantigen (TSST-1, staphylococcal enterotoxin B & C).
Types :
a. Menstrual (D/t use of high absorbency tampons).
b. Non-menstrual.
Superantigens activate 5-20% of Th cells
Cytokine storm
Toxic shock syndrome

Rx : Clindamycin (↓ses toxin production)
Rx of S. aureus infection :
1. MSSA organisms :
• TOC : β lactams.
• β lactam resistance : 90%.
• M/c mode of resistance : By penicillinase enzyme.
• Encoded by β lactamase (bla) gene on R plasmid.
2. MRSA organisms :
D/t SCCmec (mec A gene) → Encodes altered transpeptidase (PBP2a).
Diagnosis : Nucleic acid amplification test (NAAT) - Gold standard.
Rx :
• DOC : Vancomycin.
• Other drugs :
a. Daptomycin.
b. Clindamycin/Cotrimoxazole/Linezolid for skin & soft tissue infections (SSTI).
3. Vancomycin resistance : ----- Active space -----
Vancomycin Intermediate Vancomycin Resistant

S. aureus (VISA) S. aureus (VRSA)
MIC 4-8 microgram/ml ≥ 16 microgram/ml
Chromosomal mutation → Van a gene encoded on
Resistance
Thickened cell wall with ↑sed no. transposon/plasmid → Altered
method
of unlinked d-ala-d-ala terminals. drug target (d-ala-d-lactate).
Note : Vancomycin resistance cannot be determined by disc-diffusion method,
instead MIC (Minimum inhibitory concentration) is used.
S. aureus carrier :
M/C site : Nose, oropharynx.
Elimination of carriers :
• Nose : Mupirocin, bacitracin & neomycin ointments.
• Areas other than nose : Chlorhexidine soaps/washes.
• Complicated cases : Antibiotics + Rifampicin x 1 week.
M/C mode of spread in hospitals : Hands of health care personnel.
Most important step in prevention & spread in hospitals : By hand hygiene.
Maintain contact precautions while interacting with MRSA patients.
Streptococcaceae 00:24:22
Catalase -ve; Oxidase -ve.

Fastidious (Requires blood).
Brown’s classification of Streptococci :
Streptococci
α hemolytic β hemolytic γ hemolytic
Viridans Pneumococcus Enterococcus Non

group enterococcal
(VGS) group D
Lancefield serogroups based on
carbohydrate Ag on cell wall
Group A βHS Group B βHS Group C, D,...., V βHS

S. pyogenes S. agalactiae
18 02 Microbiology

S. viridans vs pneumococcus :
Viridans group Pneumococcus
Morphology Chains Pairs
Bile solubility Insoluble Soluble
Optochin Resistant Sensitive
Inulin fermentation - +
Viridans group streptococci :
Part of normal flora; Have low virulence.
S. sanguis : M/c cause of late prosthetic valve endocarditis.
S. mutans : M/c cause of dental caries.
Pneumococcus :
Most virulent : Serotype 3.
Produce draughtsman colonies (D/t autolytic enzymes).
Virulence factors :
1. Capsule (Most important).
2. Pneumolysin (O2 labile hemolysin).
3. Autolysin (Amidase).
4. IgA 1 protease.
Infections caused :
• Invasive infections in immunocompromised pts : Like meningitis, pneumonia,
endocarditis, bacteriemia etc.,
• Non invasive infection : Local infections like otitis media, sinusitis, acute
exacerbation of COPD.
• S. pneumoniae is the m/c cause of
a. Meningitis overall.
b. Meningitis in elderly individuals.
c. Community acquired pneumonia.
d. Otitis media & sinusitis.
Treatment : β lactams.
Draughtsman colonies Gram stain of CSF specimen showing G+ve

produced by pneumococci cocci in pairs suggestive of pneumococci
Note : Rapid urine antigen tests +ve in pneumococcal pneumonia & legionnaire’s disease. ----- Active space -----
Pneumococcal vaccines :
Type specific subunit vaccines based on capsular antigen.
2 types :
1. PPSV (Pneumococcal polysaccharide vaccine)/Pneumovax : Contains 23
capsular antigens, but not to be given in babies <2 years.
2. PCV (Pneumococcal conjugate vaccine)/Prevnar : 10 to 20 capsular antigens
conjugated with a carrier protein CRM197 to increase the antigenicity.
β hemolytic streptococci :
β hemolytic streptococci
S. pyogenes S. agalactiae
Group A carbohydrate Ag Group B carbohydrate Ag
Bacitracin sensitive Bacitracin resistant
Ribose fermentation -ve Ribose fermentation -ve
Both cAMP and hippurate -ve Both cAMP and hippurate +ve
S. agalactiae :
Normal flora in GIT & GU tract.
Common cause for neonatal meningitis & neonatal sepsis.
Management :
1. Puerperal sepsis : β lactam antibiotics (Penicillin/Ampicillin).
2. Prevention of neonatal infection : Rectal & lower vaginal swab at 35-37 weeks
of pregnancy → If +ve → Intrapartum penicillin prophylaxis.
S. pyogenes :
Hyaluronic acid capsule; Only 1 antigenic type.
Griffith typing : According to M protein (Present in cell wall).
Virulence factor :
1. Streptolysin O (Oxygen labile) : Antigenic (ASO is used in rheumatic fever
diagnosis).
2. Streptolysin S (Oxygen stable) : Non antigenic.
3. Streptodornase/DNAse (A-D) :
Protects bacteria from NETs (Neutrophil extracellular traps).
DNAse B is most antigenic.
Anti-DNAse B Ab titer is used in retrospective diagnosis of PSGN.
20 02 Microbiology

4. Streptokinase (Fibrinolysin) : Helps in the spread of infection by lysis of fibrin
(Fibrinolysin produced by Strep. equisimilis is used in the lysis of clots in MI).
5. Superantigens (Erythrogenic/pyrogenic toxins A, B & C).
Infections :
M/C cause of bacterial sore throat, superficial lymphangitis.
Also causes impetigo, SSTI, cellulitis, necrotizing fascitis.
Diagnosis :
Specimen : Throat swab.
Transport medium : Pike’s medium.
Gold standard : Culture on crystal violet blood agar (CVBA).
Colony morphology : Matt or mucoid colonies (Avirulent strains : Glossy colonies)
Biochemical tests :
• Bacitracin sensitive.
• Ribose +ve.
• PYR +ve.
Rapid tests : Based on immuno chromatography.
Non suppurative complications :
• Acute Rheumatic fever.
Pathogenesis : Molecular mimicry of M protein.
• Glomerulonephritis.
Acute Rheumatic fever Glomerulonephritis

Only follows pharyngitis Follows pharyngitis or pyoderma
Nephretogenic M types :
Rhegmatogenous M types :
M-1, -4, -12 & -25 after pharyngitis;
M-1, -3, -5, -6, -14, -18, -19
M-2, -9, -55, -57, -59, -60 & -61 after pyoderma.
Diagnosis :
• Throat culture positive.
• ASO ↑
Toxin mediated syndromes :

Caused d/t pyrogenic/erythrogenic/scarletiniform toxins species A to C.
1. Scarlet fever : Fever, sore throat, cervical lymphadenopathy, strawberry
tongue, scarletiniform rash, circumoral pallor & pastia lines in axilla & groin.
2. Toxic shock syndrome :
• D/t cytokine storm.
• Treated by Penicillin + Clindamycin + pooled IVIGs.

γ hemolytic Streptococcus : ----- Active space -----

They have group D carbohydrate antigen in the cell wall.
Non- Enterococcal group D Enterococcus
Bile resistant (Ability to grow in 40% Bile)
Aesculin +ve
No growth in 6.5% salt Grows in 6.5% salt
PYR -ve PYR +ve
Enterococcus :
Gut flora & environmental saprophyte.
Causes : UTI, bacteremia, infective endocarditis, pelvic infections.
80-90% : E. faecalis; 5-10% : E. faecium.
Treatment of Enterococcal infection :
• Ampicillin + Gentamicin combination : To overcome resistance to aminoglycosides
and cephalosporins.
• Vancomycin : To treat highly resistant strains of enterococci.
Some strains of E. faecium are resistant to Vancomycin (Altered cell wall
precursor molecules d/t van a, b,c genes).
Non-enterococcal group D :
Can cause septicemia/endocarditis in cases of colorectal carcinoma.
Colorectal carcinoma : D/t S. equinus & S. gallolyticus
Gram Positive Bacilli 00:47:16
Gram positive bacilli
Not gram stained Gram stained
Non branching Branching

Mycobacterium
Anaerobe Aerobe Anaerobe Aerobe
Clostridium Corynebacterium Actinomyces Nocardia

Bacillus
Listeria
Erysipelothrix
Spore forming bacteria : Clostridium & Bacillus

22 02 Microbiology
----- Active space ----- Mycobacterium :

• Thick peptidoglycan + lipid layers → Does not let the gram stain through.
• Acid fast.
Atypical mycobacteria (ATM) :

• Aka Non-tubercle/Mott (Mycobacteria other than tubercle bacillus)/
Environmental Mycobacteria.
• Acid fast.
• Not alcohol fast.
• Differentiated from mycobacteria : Niacin test -ve & arylsulfatase test +ve.
• On LJ medium, most of them grow slowly (2-8 weeks).
Runyon classification :
Based on their growth in LJ medium, they are classified into
1. Photochromogens (Pigment produced when exposed to light) : “MASK”
Marinum, Asiatum, Simiae, Kansasii.
2. Scotochromogens (Pigment produced both in light & dark) : Szulgai,
Scrofulaceum, Gordonae.
3. Non-photochromogens (No pigment produced) : MAC (Mycobacterium avium
intracellulare complex), Ulcerans, Xenopi.
4. Rapid growers (Colonies formed within 5-7 days) : “AFMC” Abscessus,
Fortuitum, Mucogenicum, Chelonei.
Note :
Mycolactone : An exotoxin produced by Mycobacterium ulcerans.
M. indicus pranii species (Mycobacterium W) is used for leprosy vaccine.
Infections caused by ATM :

Reservoirs of ATM are soil & water.
ATM are low virulent organisms which infect immuno-compromised individuals.
1. SSTI :
• Buruli ulcer caused by M. ulcerans.
• Swimming pool granuloma caused by M. marinum (Fish tubercle bacillus).
• Post- traumatic/Surgical abscess is most commonly caused by Rapid
growers.
2. Lymphadeinitis : MAC > M. scrofulaceum.
3. Pulmonary infection : MAC > m. kansasii.

Mycobacterium Tuberculosis Complex : ----- Active space -----

• Acid fast and Alcohol fast.
• Niacin test +ve & Arylsulfatase test -ve.
• Strict Aerobe.
Diagnosis :
1. Microscopy :
Most rapid method.
Stain : Ziehl–Neelsen, Kinyoun.
Sensitivity : 10,000/ml of specimen.
Fluorescent stains : Auramine O/Rhodamine B (↑sed sensitivity).
2. Culture :
Gold standard.
Good sensitivity : 10-100/ml.
Generation time : 14-15 hrs.
Manual methods :
• Solid medium : LJ medium, Dorset egg medium (3-8 weeks).
• Liquid medium : Middle brook medium (5-10 days).
Automated method : Uses liquid media.
• Mycobacterium growth indicator tube (MGIT).
• BacT-Alert (Automated colorimetric method).
3. Molecular methods :
• Hybridisation probes.
• NAAT : PCR/Real time PCR (Xpert MTB-Rif assay/Xpert MTB-Rif Ultra).
Genes amplified : IS6110 > IS1081.
Counter stain : Methylene blue LJ medium containing Rough, tough & buff colonies
egg & malachite green
(Selecting agent)

24 02 Microbiology

4. Susceptibility tests :
a. Conventional methods : Takes long time.
LJ medium with incorporated anti tubercular drugs.
b. Microscopic observation of drug susceptibility.
c. Automated method : MGIT/BacT-Alert with drugs added.
d. Molecular tests : Looks for specific gene mutations (rpo B/inh A).
5. Lipoarabinomannan (LAM) in urine : Used in detection of active TB in HIV pts.
6. Exposure to MTB :
• Mantoux/tuberculin test : Purified protein derivative injected intradermally
in forearm → Induration is measured.
• Interferon γ release assay (QuantiFERON gold test, TB spot test) : CFP-10,
ESAT-6, TB 7.7 are the antigens added to whole blood.
BCG vaccine :
Live attenuated vaccine of M. bovis strain.
Danish 1331 strain.
Spore Forming Bacteria 01:01:30
Bacillus Clostridium
Forms Non-bulging spores. Forms Bulging spores.
Grows in soil & culture. Grows in soil, cultures & body.
Subterminal :
• C. perfringens
• C. botulinum
Central/ subterminal.
Terminal :
• C. tetani
• C. difficile
Clostridium :
Strict anaerobe.
Culture : Robertson cooked meat broth.
Lacks catalase, oxidase, and superoxide dismutase.
C. botulinum :
Botulinum toxin :
• Category A bioterrorism agent.
• Releases neurotoxin on cell lysis (Most potent exotoxin) which is heat labile.
• 8 antigenic types : A, B, E, F causes human disease. ----- Active space -----

• Botulinum toxin proteolyzes SNARE proteins (Synaptobrevin/syntaxin/
SNAP-25) → Inhibit Ach release at peripheral nerve terminals.
Food Botulism :
Preserved/canned food containing preformed toxin.
• Presents with CN deficits, followed by descending symmetric flaccid
paralysis.
• Fever & sensory deficits are absent.
• Infant botulism (floppy baby syndrome) :
Occurs mostly with honey being contaminated with spores.
Presents with GI symptoms (constipation).
• Rare type of botulism involves contamination of wound with spores
Diagnosis : Serum assay (NAAT & ELISA based tests).
Treatment :
• Equine antitoxin : Used in adults.
• Human antitoxin : Used in infant botulism.
C. tetani :
Motile, non-capsulated, drumstick morphology d/t terminal spores.
Shows swarming motility (Filde’s technique of isolation done).
Tetanospasmin :
• Neurotoxin.
• Proteolyses synaptobrevin → Presynaptic inhibition of GABA & glycine in brain
stem & spinal cord → Uncontrolled excitation → Spastic paralysis.
Risk factor : Traumatic entry of spores.
IP : 6 to 12 days.
C/F :
• M/c : Trismus (Lock jaw).
• Risus sardonicus (D/t spasm of orbicularis oris).
• Tetanic spasms.
• Descending symmetric spastic paralysis & opisthotonus.
Treatment :
• Human tetanus immunoglobin (HTIG).
• Age appropriate tetanus toxoid.
• Supportive Mx and I/v Metronidazole.

26 02 Microbiology

C. perfringens :
Non motile, capsulated & subterminal spores. Has ability to grow at 450 C.
Special features :
• Stormy fermentation at litmus milk.
• Target haemolysis.
• Reverse CAMP test +ve.
Major toxins :
• α, β, ξ, and iota toxin.
• α toxin :
Has lecithinase activity detected by Nagler reaction.
Lecithinase causes membrane damage, platelet aggregation and
intravascular hemolysis.
Type A C. perfringens : Causes gas gangrene and food poisoning.
Nagler’s reaction done on egg yolk agar
Gas gangrene :
Occurs when spores enter traumatically (IP : 1-3 days).
C/F :
• Pain, discharge, edema, crepitation.
• Toxemia, shock (D/t rapid spread to underlying tissues).
Rx : CT/MRI → Emergency surgical resection & penicillin with clindamycin.
Food poisoning :
Meat contaminated with heat resistant spores (IP : 8-24 hrs).
Enterotoxin :
• Coded by CPE gene.
• Formed in the GI tract & causes pore formation in enterocytes.
Diagnosis : Detection of CPE gene in stool samples by ELISA and amplification
techniques.
Rx : Symptomatic treatment.
C. difficile : ----- Active space -----

Motile, non capsulated, normal GI flora.
Terminal oval spores which gives tennis racquet appearance.
Toxins :
6-8% strains produce Toxin A (enterotoxin) and Toxin B (cytotoxin).
Disruption of actin cytoskeleton → Apoptosis.
H/o taking antibiotics like Ampicillin, Amoxycillin, Clindamycin or Fluoroquinolones
→ Overgrowth of toxigenic C.difficile.
IP variable.
C/F :
• Watery diarrhea + abdominal cramps + fever.
• Severe cases : Toxic megacolon or paralytic ileus (Need parenteral antibiotics).
Diagnosis
In stool
Organism Toxin Colonoscopy
• Culture : CCFA medium. • PCR.
• Glutamate dehydrogenase • Cytotoxicity Pseudomembrane
(GDH) assay. assay. (severe disease)
Treatment :
• Mild - moderate cases :
Oral Fidaxomicin > oral Vancomycin
• Severe cases :
Parenteral Vancomycin & Metronidazole.
Pseudomembrane

28 02 Microbiology

Branching G+ve Rods 01:15:22
Actinomyces Nocardia
Staining Non-acid fast Partial acid fast
O2
Strict anaerobes. Aerobe.
requirement
Normal commensals in human GI, GU
Habitat Soil saprophyte.
flora.
Exogenous :
Mode of
Endogenous. • Trauma (M/C sp. : N. braziliensis).
infection
• Inhalation.
Manifests as chronic/subacute • Cutaneous : Cellulitis, nodular
granulomatous disease with hard lymphangitis (Sporotrichoid lesions),
nodular appearance and sometimes Actinomycetoma → Disseminated
with sinuses. nocardiosis (in immunocompromised).
• M/c : Cervicofacial involvement • Inhaled : Pleuropulmonary nocardiosis
C/F
(Lumpy jaw : Follows Sx/ (In immunosuppressed pts).
poor dental hygiene/radiotherapy).
• Thoracic.
• Abdominal.
• Pelvic : A/w IUCD.
Paraffin bait technique is
Diagnosis Forms molar tooth colonies. used to isolate from soil.
Pigment +/-
Sulfonamides (TMP-SMX) +/-
Rx Penicillin
Amikacin & carbapenem.
Actinomycosis :
Lumpy jaw
Sulfur granules
Images
Sunray appearance on histology/

Splendor hoepplei phenomenon
Branching G+ve rods

Corynebacterium diphtheriae 01:21:34 ----- Active space -----
Non motile, non capsulated, non sporing, has chinese letter pattern.
Contain metachromatic/volutin/polymetaphosphate/babes-ernst granules →
Stained by Albert/Neisser/Ponder stain.
Albert stain :
• Toluidine blue (Stains granules).
• Malachite green (Stains cytoplasm).
• Iodine (Mordant).
Diphtheria toxin :
• Mediated by β phage.
• Toxin gets produced only if iron values get low to a certain level.
• MOA : Inhibition of protein synthesis Via ADP ribosylation of EF-2.
• Diphtheria toxoid is prepared from Park William 8 strain.
Transmission : Via droplets; IP : 3-4 days.
C/F :
• Faucial diphtheria (M/c) : Fever, sore throat, pseudomembrane over tonsil.
• Laryngeal type : Dangerous & requires immediate tracheostomy.
• Complications :
1. Myocarditis (M/C complication & M/C cause of death in diphtheria).
2. Neurological deficits :
- Palatal and ciliary nerve palsies.
- D/t demyelination of neurons.
- Patient recovers spontaneously.
Diagnosis :
• Gram stain (Club shaped rods) & Albert stain.
• Culture : Loeffler’s serum slope (Forms colonies in 6-8 hrs).
• Selective media : Potassium tellurite media like Tinsdale & Mcleod’s media.
• Elek’s gel precipitation test (or ELISA) : To demonstrate toxin formation in
colonies.
Rx :
Anti diphtheria toxin Antibiotics
Based on disease duration + Procaine penicillin (or)
• <3 days : 20-60k U x 2 weeks
IM/IV Erythromycin
• >3 days : 80-120k U

30 02 Microbiology
----- Active space ----- Prophylaxis for contacts : Penicillin/Erythromycin ± DTaP/dTaP (Toxoid).
Schik test : Test for susceptibility by seeing induration after I/d inoculation.
Gram stain showing G+ve rods in Albert stain showing green rods
cuneiform arrangement with purple granules
Cutaneous diphtheria
Bull neck of severe

Faucial diphtheria faucial diphtheria
Filter paper with anti-DT
Negative control
Test isolate
Positive control
Precipitin lines
Elek’s gel precipitation test : Ouchterlony lines
Bacillus 01:26:48
B. cereus :
Part of anthracoids.
MYPA medium : Selective medium.
2 types of food poisoning :
1. Emetic type : Chinese rice (Preformed cereulide → Vagal stimulation). IP <6 hrs.
2. Diarrheal type : Meat with spores, (Nhe toxin : Formed in GI tract). IP 8- 16 hrs.
Both are self limiting.
B. anthracis : ----- Active space -----

• Category A bioterrorism agent.
• Medusa head colonies.
• Bamboo stick appearance.
• Non- motile, capsulated.
• Not hemolytic on blood agar.
• Inverted fir tree pattern in gelatin liquefaction.
• Susceptible to γ phage.
Virulence factors :
1. Polypeptide capsule (Polymer of D-glutamate) demonstrated by M’Fadyean
reaction.
2. Anthrax toxin (Protective antigen, edema factor & lethal factor) → Acts by
increasing cAMP by calmodulin mediated mechanism.
Selective medium : PLET medium.
Infective form : Spores, if acquired.
• Trauma : Hide porters disease/ malignant pustule → Septicemia (rare).
Painless ulcer + surrounding edema → Heals with eschar.
• Inhalational anthrax : Aka wool sorters disease (Highly fatal mediastinitis).
• Intestinal anthrax : Rare type of hemorrhagic enteritis.
Diagnosis : Microscopy, culture (Gold std), Ag detection, PCR, serology.
Rx :
• For Cutaneous anthrax : PO Ciproflox/Doxycycline
• For other types : IP Fluoroquinolones + β lactam + Clindamycin/Rifampicin/
Linezolid x 2-3 weeks → PO Cipro/Doxy for 60 days + AVA (0, 2, 4 weeks).
Inhalational anthrax with Eschar

mediastinal widening DD : Brown recluse spider bite, cutaneous anthrax,
spotted fever rickettsiosis, scrub typhus

32 02 Microbiology
----- Active space ----- Listeria monocytogenes 01:32:07
Unique G+ve bacterium which has cell wall with endotoxin in it.
Differential motility :
• Non- motile at 370 C.
• Motile at room temperature (Tumbling/End on end motility).
Can grow between 2 to 45°C, so cold enrichment is done.
cAMP test +ve.
PALCAM : Selective medium.
Present in soil, water & sewage.
Acquired by refrigerated foods, unpasteurized milk & milk products.
C/F :
• Mostly asymptomatic.
• Gastroenteritis in some.
• Meningitis in elderly and immunodeficient individuals.
• In pregnant females, 17-fold risk of bacteremia → Chorio-amnionitis
→ Abortion, still-births, neonatal deaths & granulomatous infantiseptica
(Multiple pustules & visceral abscesses in the newborn).
• In some pregnant females, the bacteria is carried via GI/GU tract of mother
and causes meningitis in neonates.
Doc : Ampicillin, in cases of meningitis add Gentamicin.
Listeria/S. agalactiae
S. aureus
CAMP test

Gram negative bacteria 00:00:21
Gram negative bacteria
Cocci Bacilli Coccobacilli Spirals
• Neisseria • Enterobacteriaceae • Acinetanobacter • Spirochetes

• Moraxella • Pseudomonadaceae • Brucella • Camplylobac-
• Bacteroides • Bordetella teriaceae
• Vibrionaceae • Haemophilus
• Legionella
• Mycoplasma
• Chlamydia
• Rickettssia
Note : Chronic granulomatous disease (CGD) → Mutation in NADPH oxidase
enzymes→ ↑ risk of infection by catalase +ve bacteria.
MacConkey agar:
• Mildly selective for GN bacteria.
• Indicator medium → Neutral red indicator → Differentiate lactose &
non-lactose fermentors (differential medium).
• Ingredients : PLANT → Peptone lactose agar, neutral red, sodium
taurocholate (selecting agent).

34 03 Microbiology
----- Active space ----- Lactose fermenter (LF) Non lactose fermenter (NLF)
Mnemonic : CEEK • Salmonella
• Citrobacter • Shigella
• E.coli • Proteus
• Enterobacter • Yersinia
• Klebsiella • Pseudomonas
• Shigella sonnei • Vibrio
(Late LF). Pink LF colonies • Acinetobacter Pale NLF colonies
EMB (Eosin methylen blue agar) :

• Midly selective for GN bacteria.
• Indicator medium → Eosin & methylene blue indicator → Differentiate LF &
NLF.
Enterobacteriaceae 00:05:41
Includes :
1. Shigella, 4. Proteus,
2. E. Coli, 5. Salmonella.
3. klebsiella, 6. Yersinia.
Common features :
Physical Biochemical Culture
• Motile with peritrichous flagella except: • Catalase +ve MacConkey
a. Klebsiella (except S. dys- agar & EMB
b. Shigella enteriae type 1). agar : LF/
c. Yersinia pestis • Oxidase -ve. NLF
d. Salmonella gallinarum pullorum • Nitrate reduc-
e. Atypical E.Coli. tase +ve.
• Capsule ± • Ferment sugar
• Facultative anaerobes, non fastidious to produce acid
only/ acid + gas.
1. Shigella :
• NLF except S. sonnei (late LF).
• Mannitol fermenter except S. dysenteriae.

• Virulence plasmid encodes invasive plasmid Ag/IPA proteins (VMA) & type 3 ----- Active space -----
secretion system → Helps in basolateral invasion of colon and rectum.
• S. dysenteriae type 1 → Shiga toxin
Necrosis of Receptor : Gb3 Cleaves 60s Hemolytic uraemic Local ischemia

endothelial ribosome & inhibit syndrome (HUS).
cells protein synthesis (M/C cause in india :
S. dysentriae type 1).
M/c cause of shigellosis in India→ S.flexneri & in world → S. sonnei.

Most communicable bacterial diarrhea : Shigellosis (d/t small infective dose).
Clinical features : Bacillary dysentery → Fever, abdominal pain, blood & mucus in
stools.
Diagnosis
Transport medium Culture Sereny test Lab diagnosis
• Buffered • Enrichment medium : Keratoconjuctivitis • Demonstration

glycerol Selenite F broth. on inoculation into on cell lines.
medium • Selective broth : guinea pig. • ELISA & PCR.
• Cary blair a. DCA (deoxycholate
medium citrate broth).
b. XLD (Xylose lysone
deoxycholate agar)
c. SSA (Salmonella
shigella agar).
d. HEA (Hektoen enteric
agar).
Complications : Toxic megacolon, HUS, reactive arthritis (S. flexneri).
Mx :
• Fluid & electrolytes
• 3rd generation cephalosporins : Severe disease, extremes of age,
immunocompromised.
Causes of invasive diarrhea :
• Vibrio parahaemolyticus.
• Balantidium coli.
• E. histolytica.
36 03 Microbiology

• Shigella.
• EIEC (Enteroinvasive E.Coli) & EHEC.
• Non-typhoidal salmonella.
• Campylobacter.
• Yersinia enterocolitica
2. E.coli
• Motile (Exception : Atypical E. Coli).
• Capsular K Ag, somatic O Ag & flagellar H Ag serotype.
• LF.
• Biochemical : Produce acid + gas & indole+ve.
Clinical features
UTI Neonatal meningitis Diarrhea Intrabdominal Hospital

• Uropathogenic abscess aquired
E. Coli (UPEC). pneumonia
• P pili & type 1 pili
(Adhere to urothelial mucosa).
Diarrheagenic Features Virulence factors
E. Coli
1. Enterotoxigenic • M/C cause • Colonisation factor antigens
(ETEC) of bacterial & • Heat labile toxin : ↑cAMP.
traveler’s diarrhea. • Heat stable toxin : ↑cGMP.
• Infective dose > 109.
2. Enteroinvasive • Atypical E. Coli→ • VMA/IPA protein.
(EIEC) Nonmotile, NLF, • Type 3 secretion system
anaerogenic.
• Bacillary dysentery.
• Very ↑ infective
dose.

3. Enteropathogenic • MCC of infantile • EPEC adherence factor ----- Active space -----
(EPEC) bacterial diarrhea. plasmid (Bundle forming pili)

• Locus of enterocyte
effacement plasmid :
Attachment effacement
effect.
3. • MCC of persistent • Aggregative adherence

Enteroaggregative diarrhea fimbriae (AAF)
(EAEC) : • Causes traveller’s • EAST-1 : Enteroaggreg Active
diarrhea. Stable Toxin) : ↑cGMP.
• Culture : Hep 2
adherence shows
stacked brick
adherence.
4. • Zoonoses. • Shiga-like toxin/ verotoxin
Enterohemorrhagic • Contaminated (same action as Shiga toxin).
(EHEC) : beef/hamburgers. • Locus of enterocyte
• Small infective dose. effacement plasmid.
• E. Coli O157 : H7 →
Severe colitis & MCC
of HUS in world.
• Sorbitol McConkey
agar : Pale colonies
(Non-sorbitol
fermenters).
3. Klebsiella pneumoniae/Friedlander’s bacillus

• Capsulated, non-motile.
• LF : Mucoid colonies on McConkey agar.
• Indole -ve.
• Hypermucoviscous strain of klebsiella :
a. Thick capsule → String Test +ve
b. M/C cause of intra-abdominal abscess.
Belongs to ESKAPE group of MDR (multidrug-resistant) organisms :

E : Enterococcus faecium.

38 03 Microbiology

S : S. aureus.
K : klebsiella pneumoniae.
A : Acinetobacter baumannii.
P : Pseudomonas aeruginosa.
E : Enterobacter.
Clinical features :
1. Friedlander’s pneumonia :
• Seen in DM pts and alcoholics.
• Red currant jelly sputum.
• Upper lobe consolidation with bulging fissure sign.
2. UTI, neonatal septicaemia, intraabdominal abscess
3. Rhinoscleroma :
• Caused by K. pneumoniae rhinoscleromatis.
• Nodules on upper respiratory tract, hebra nose.
• Rx : Tetracyclines, streptomycin, fluoroquinolones.
Mikulicz cells on biopsy Nodules in URT Hebra nose

4. Ozaena/atrophic rhinitis :
• Caused by K. pneumoniae ozaenae.
• Crusting and foul smelling discharge with merciful anosmia.
• Rx : Tetracyclin/Fluoroquinolones.
5. Donovanosis/granuloma inguinale :
• Caused by K. granulomatis.
• Painless beefy red genital ulcer, rolled edges, bleeds, no lymphadenopathy.
• Pseudobubo : Inguinal granuloma with lymphadenopathy -ve.
• Rx : Azithromycin for atleast 3 weeks.
Genital ulcers in donovanosis Donovan bodies seen in pund cells

3. Proteus : ----- Active space -----

• Motile.
• PPA (Phenyl pyruvic acid) test +ve.
• Urease +ve.
• NLF, Swarming on medium & fishy smell.
• Diene’s phenomenon : Swarming merge
Swarming Swarming seperated
if same strains of proteus & separated by by zone.
a zone if different strains.
Clinical features :
• UTI
• Recurrent infection → leads to persistent alkalization of urine d/t urease
enzyme → formation of struvite stone.
UTI :
GN GP
E. coli (M/c) S. saprophyticus (UTI in sexually active females)
Klebsiella S. aureus
Enterobacter Entercoccus
Citrobacter Candida
Proteus
Pseudomonos
Causes of UTI Special features

E. coli Indole +ve
Klebsiella Indole -ve
Urease +ve
Enterobacter Indole -ve
Citrobacter Indole -ve
Proteus PPA +ve
Urease +ve
Swarming +ve
Pseudomonos Grows at 420 C
Oxidase +ve

40 03 Microbiology
----- Active space ----- Diagnosis
Microscopy Dipstick Culture :

Gold standard.
• wet mount > 10 WBC/hpf • Leucocyte esterase test :
• Gram stain > 1/oil Presence of pus cells.
immersion field. • Griess nitrate test :
Detect nitrite reducing
bacteria.
• CLED (Cystine lactose electrolyte deficient) medium preferred over

MacConkey for diagnosis of UTI.
• CLED allows growth of both GP and GN (Non-selective medium).
• MacConkey is selective to GN.
5. Salmonella :
Motile, non-capsulated except typhi, {paratyphi & dublin → Polysaccharide
capsule (Vi antigen)}.
Somatic O Ag, polysaccharide capsular Vi Ag, flagellar H Ag serotypes.
NLF.
Culture :
• Transport medium : Cary Blair medium.
• Enrichment medium : Selenite F broth, tetrathionate broth.
• Selective medium : DCA, XLD, SSA, HEA.
• Wilson Blair medium (best) : Black colonies d/t H2S.
Clinical features :
• Enteric fever : Typhoid & paratyphoid fever.
• Caused by S. typhi, paratyphi A, B & C.
• Prolonged step ladder pyrexia.
• Faget sign (fever with bradycardia).
• Rose spots.
Complications :
• Gl bleeding (M/c).
• Perforation.
Diagnosis :
• Gold standard : Culture
• Blood culture : +ve in 1st week on Castaneda biphasic medium.
• Stool & urine culture : +ve at the end of 2nd/3rd week. ----- Active space -----
• Bone marrow culture : Most sensitive (+ve even after start of Rx).
Widal test :
• Tube agglutination test, detect O & H antibodies.
• O antibodies → Recent infection.
• H antibodies → To detect serotype.
Rx :
Empirical → Ceftriaxone/azithromycin .
Definitive : Ofloxacin/ciplox for 5 to 14 days.
Non typhoidal salmonella :

• S. typhimurium & S. enteritidis → Invasive diarrhea (zoonotic) → self
resolving.
• S. cholerasuis & S. dublins → Septicemia seen in extremes of age,
immunocompromised.
• Rx : 3rd generation cephalosporins.
Typhoid vaccines :
• Typhoid Vi : Contains Vi polysaccharide of S. typhi.
Given S/c
• Conjugated Vi polysaccharide vaccine : Conjugated with
> 2 yrs of age.
TI/nontoxic exotoxin A of P. aeruginosa.
• Oral live atenuated typhoral vacine : Ty21a for children > 6 years.
6. Yersinia pestis
Category A bioterrorism agent (BSL-3 organism).
Virulence factors : F1 capsular antigen.
NLF, non-motile.
Clinical features :
Plague
• Vector : Xenopsylla cheopis (rat flea) & reservoir → rats
• Types :
a. Bubonic (M/c) : Transmitted by bite of rat flea, painful enlarged LN.
b. Pneumonic : d/t inhalation of aerosols causes hemorrhagic pneumonia.
c. Septicemic.
Diagnosis :
• Wayson stain : Safety pin appearance.
Culture :
• Selective medium → CIN (cefsulodin irgasan
novobiocin) agar & ghee broth shows stalactite growth.
Safety pin appearance

42 03 Microbiology

• Grows at 2 to 45°C → Cold enrichment → Other safety pin
Isolation by incubating in refrigerator (also appearance :
shown by listeria monocytogenes). • Klebsiella granulomatis.
• Haemophilus ducreyi,
• Detection of F1 antigen & NAAT to detect gene.
• Burkholderia mallei &
Rx : pseudomallei.
• Streptomycin, gentamicin, chioramphenicol.
• Prophylaxis : Doxycycline, fluoroquinolones.
Yersinia enterocolitica & Yersinia pseudotuberculosis :

• Differential motility : Motile at room temperature & non-motile at 37 degree
celsius (also shown by listeria).
• Zoonotic.
• Reservoir : Cattle, deer, pigs & birds.
• Causes :
a. Yersiniosis.
b. Mesenteric adenitis & terminal ileitis : Pseudoappendicitis.
c. Y. enterocolitica : Self-limited diarrhea.
d. Y. pseudotuberculosis : Septicemia in iron overload states (also caused
by Listeria & vibrio vulnificus).
• Rx : Fluoroquinolones, 3rd generation cephalosporins.
Vibrio 00:50:53
Comma shaped, darting/shooting star motility with monotrichous flagella.

Alkaliphiles (pH 8.2 to 8.4)
Halophiles (grows at 1% salt concentration) except v.cholerae & v.mimicus.
Reservoir : Sea water.
Catalase, oxidase +ve.
Cholera red reaction +ve.
String test +ve (string on addition of bile salt).
NLF
TCBS agar: Differentiate sucrose fermenting v. cholerae & non-sucrose
fermenting v. parahaemolyticus
Gardner & venkatraman classification :

• Group A : v. cholerae with H antigen.
• Group B : All other vibrios.

1. V. cholerae ----- Active space -----

Cholera toxin
Produced by Bacteriophage Receptor : GM1 ADP ribosylation →

V.cholerae 01 (El mediated ganglioside on ileal ↑ adenyl cyclase
Tor & classical) & mucous cells activity → ↑cAMP.
V.cholerae 0139
(Bengal strain)
Other virulence factors :

Toxin coregulated pilus, zonula occludens toxin (disrupt tight junctions b/w
enterocytes).
Clinical features :
• Risk factors : Achlorhydria, H/o gastrectomy and blood group O.
• Cholera : Rice water stools and metabolic acidosis.
Diagnosis:
• Transport medium : Venkatraman Ramakrishnan & Cary Blair.
• Wet mount : Fish in stream appearance.
• Enrichment medium : Alkaline peptone water.
• Selective medium : Yellow colonies on TCBS.
• Serotyping to detect serotypes.
Mx :
• Fluid & electrolyte balance.
• Moderate/severe dehydration : Doxycycline/azithromycin.
Cholera vaccines :
Dukoral Shanchol/Euvichol Orochol/
plus Vaxchora
Route Oral Oral Oral
Type Killed v. cholera O1 + Killed v. cholerae O1 + Live attenuated
B subunit of cholera v. cholerae O139 strain of v.
toxin cholerae 01 (CVD
103 HgR)
Age group ≥2 yrs ≥1 yr 2 to 64 yrs

44 03 Microbiology

Group B vibrios :
V. parahemolyticus V. vulnificus V. alginolyticus
• Liquid media : Monotrichous. • LF. • Grows upto 10%
• Solid media : Peritrichous. • Cellulitis following salt
• Grows upto 8% salt conc : sea water trauma. • Cellulitis following
Swarming + ve. • Seafood associated sea water
• On Wagatsuma agar : septicaemia in iron trauma.
kanagawa phenomenon overload states &
• Seafood (oysters & shellfish) diarrhea.
associated invasive diarrhea
• Rx : Doxycycline.
Pseudomonadaceae 01:02:22
Motile, strict aerobes.

Catalase & oxidase +ve
Saprophytes in moist environments.
Pseudomonadaceae
1. Strenotrophomonas 2. Pseudomonas 3. Burkholderia
• Nosocomial & opportunistic infections in

cancer & cystic fibrosis.
• Intrinsic resistance to aminogylcosides &
beta lactams including carbapenams.
• DOC : Cotrimoxazole.
2. Pseudomonas aerogenosa :
• NLF, mucoid colonies .

• Grows at 42°C.
• Pyocyanin, pyomelanin, pyorubin,
pyoverdine pigment.
• Selective medium : Cetrimide agar → Pyocyanin Pyoverdin
Fruity smell colonies.

Virulence factors : ----- Active space -----

Slime layer : Biofilm.
Toxin : Exotoxin A which Acts by ADP ribosylation of EF-2 → Protein synthesis
inhibition
Clinical features :
Corneal ulcers in contact lens users.
Swimmer’s ear.
Malignant otitis externa.
Jaccuzi syndrome (hot tub folliculitis). Malignant otitis externa.
M/c GN causing hospital acquired & ventilator associated pneumonia.
Catheter associated UTI.
In neutropenic patients : Ecthyma gangrenosum (Necrotic skin lesion) &
septicemia.
Jaccuzi syndrome (hot tub folliculitis) Ecthyma gangrenosum
ESKAPE group of bacteria.

MDR → D/t reduced entry, efflux pumps, inactivating enzymes.
Scromboid fish poisoning :

• D/t improper processed or stored fish.
• E.coli, P. aeruginosa, klebsiella, proteus & morganella in fish cutis/intestines.
• Decarboxylate histidine in fish muscles to histamine → Flushing, rash,
urticaria, palpitations, headache, dizziness, sweating

46 03 Microbiology

3. Burkholderia :
1. B. mallei 2. B pseudomallei 3. B. capecia
• Non-motile, bipolar • Bipolar staining • Selective medium :
staining/safety pin • Caused melioidosis Ashdown medium.
appearance. : Pneumonia, • Cepacia syndrome :
• Reservoir : Equines. skin lesion, Rapid loss of pulmonary
• Can cause infection in both septicaemia. function & pneumonia
humans and horses. • Latent infection in CGD & cystic fibrosis
• Rx : Ceftazidime/ & reactivate patients.
carbapenem for 2 weeks & after months • Intrinsic resistance
doxycycline/cotrimoxazole (Vietnamese time to fluoroquinolones &
for 2 to 3 months. bomb). polymyxin.
• Rx same as mallei. • Rx : Cotrimoxazole
(DOC), ceftazidime,
carbapenems,
minocycline.
Agents causing disease in cystic fibrosis & chronic granulomatous disease (CGD) :
Cystic fibrosis CGD
• S. aureus • B. cepacia
• P. aeruginosa • Nocardia
• H. influenzae • Aspergillus
• B. cepacia • S. Aureus
• Actinomycetes
Bacteriodaceae 01:13:24
GN anaerobic rods.
Non-motile, non-spore forming.
1. Bacteroides :
M/c colon flora.
M/c clinical isolate : B. fragilis.
B. fragilis :

• LPS with low endotoxic activity (Shock and DIC risk is low). ----- Active space -----
• Clinical features : Peritonitis ±, septicemia, intraabdominal abscess.
• Beta-lactamases +ve & Intrinsic resistance to vancomycin, kanamycin,
colistin.
• Rx : Carbapenems, betalactams/betalactamase inhibitors, metronidazole.
2. Prevotella. Oral & GI flora

3. Porphyromonas Causes periodontitis & aspiration pneumonia
Gram negative diplococci 01:17:59
1. Neisseria.
2. Moraxella : M. lacunata→ Angular conjunctivitis
M. catarrhalis→ Otitis media, sinusitis, COPD exacerbation.
Neisseria :
General features Biochemical Culture
• GN non motile. Catalase & oxidase • Transport media : Stuart &
• Facultative +ve. Amie’s
intracellular. • Selective medium :
• Neisseria→ Type 4 pili Modified Thayer Martin
→ Twitching motility. medium.
• Lipooligosaccharides
+ve in cell wall (also
seen in Hemophilus).
N. meningitidis N. gonorrhea
Strict aerobe Facultative anaerobe
Capsule + Capsule absent
Utilize glucose & maltose Utilize only glucose
Facultative intracellular :
(mnemonic : Some Nasty Bugs May Live FacultativeLY).
• Salmonella.
• Neisseria.
• Brucella
• M. TB.
48 03 Microbiology

• Legionella.
• Francisella.
• Listeria.
• Yersinia.
1. Meningococcus :
Lens shaped.
Capsular serotypes : A, B, C → Mostly cause disease (M/c in India : A).
W, X, Y → Cause sporadic infection.
Virulence factors :
• LOS.
• Capsule.
• Outer membrane proteins.
• Pili.
• IgA1 protease (also produced by pneumococci, H. influenzae).
Risk factors :
• C5-C9 deficiency.
• HIV.
• Asplenia.
• Hypogammaglobulinemia.
• Adolescents, military recruits, Africa travel. Non-blanching hemorrhagic
Clinical features : rash
Meningitis Meningococcemia
Non-blanching Endotoxemia with hypotension, DIC, adrenal hemorrhage
hemorrhagic rash (waterhouse-Friderichsen syndrome).
Diagnosis :
• Gram stain: GN diplococci
• CSF : ↑ PMNs, ↓ glucose, ↑ proteins.
• Culture : Blood agar, chocolate agar.
• PCR.
Rx :
• Ceftriaxone, cefotaxime, meropenem.
• Prophylaxis : Single dose of ciplox/ceftriaxone or rifampicin for 2 days.
Vaccines :
• Meningococcal conjugate vaccine-4 : Contains A, C, W, Y capsular antigens
conjugated with dT/TT/non-toxic toxin of C. diphtheria CRM197.

• Meningococcal vaccine B-4C/Meningococcal B-FHbp : Contains fimbrial ----- Active space -----
proteins.
• Given at 11 to 12 years & booster at 16 years.
2. Gonococcus :
Bean/kidney-shaped, serotyping based on outer membrane protein.
Virulence factors :
• LOS & type 4 pili both show antigenic variation.
• Outer membrane proteins (Opa,porin,RMP).
• IgA1 protease.
Clinical features :
1. In Males :
• Urethritis : Purulent discharge with dysuria.
• Epididymitis, prostatitis, seminal vesiculitis.
2. In Females :
• Cervicitis, endometritis, salpingitis, urethritis.
• PID.
• Fitz-Hugh-Curtis syndrome (Perihepatitis).
• Tubo-ovarian abscess.
• Vaginitis in pre-pubertal & post-menopausal.
3. Pharyngitis, proctitis, conjunctivitis.
4. Ophthalmia neonatorum.
5. Gonococcemia :
• Risk factors : C5 to C9 deficiency, menstruation
• Dermatitis, large joint Arthritis, Tenosynovitis, Meningitis, Endocarditis
(mnemonic : DATe ME).
Rx : Ceftriaxone + azithromycin (if associated chlamydial infection).

50 04 Microbiology
----- Active space ----- MICROBIOLOGY REVISION 4
Gram negative bacteria : Coccobacilli 00:00:35
All Gram -ve Coccobacilli are non motile except Legionella & Bartonella.
1. Acinetobacter 00:02:10
Features :
• Non motile. • Non-lactose fermenting colonies.
• Aerobic organism. • Catalase + ve.
• Non fastidious. • Oxidase -ve.
Note : Acinetobacter baumani → Most commonly associated with human
infection.
Diseases caused : Mainly nosocomial.

1. Ventilator associated pneumonia. 3. Wound infection.
2. Blood stream infection. 4. Meningitis following neurosurgery.
Resistance :
Mechanism of resistance Resistant antibiotics
Penicillin.
Produce b lactamases intrinsically
1 & 2 gen. cephalosporins.
st nd
Plasmid : Extended spectrum b lactamases 3rd & 4th gen. cephalosporins.
Treatment :
Sulbactam and Carbapenems.
Certain Carbapenem Resistant Acinetobacter Baumani (CRAB) :
• Polymyxins.
• Sulbactam + Durlobactam.
• Tetracycline.

2. Bordetella pertussis 00:05:00 ----- Active space -----

Causes pertussis/Whooping cough/100 day cough.
Features :
• Non-motile. • Non-antigenic capsule.
• Strict aerobe. • Fastidious.
Note :
Strict aerobic bacteria :
• M. tuberculosis. • Bordetella.
• Brucella. • P. aeruginosa.
• Nocardia.
Culture :
• Media : Bordet Gengou medium & Regan Lowe medium.
• Colonies : Bisected pearl colonies/Mercury drop colonies.
Toxins :
1. Pertussis toxin :
• Helps in adheshion.
• Inhibits phagocytic function.
2. Adenyl cyclase toxin : ↑ cAMP.
Transmission : Droplet transmission.

Incubation period : 7-10 days.
Pathogenesis :
1. Catarrhal stage : 3. Convalescent stage.
• Most infectious stage.
• Antibiotics halt progression of disease.
2. Paroxysmal stage :
• Repetitive bouts of cough.
• Post-tussive vomiting, inspiratory whoop.
• Antibiotics ↓ infectivity.
Complications :
1. Surgical emphysema. 3. Hernia.
2. Pneumonia (M/C). 4. Fracture ribs.
52 04 Microbiology

Investigations :
• Lab findings : Lymphocytosis & hypoglycemia.
• Specimen : Nasopharyngeal Aspirate ( Best) > Nasopharyngeal swab > Cough
on the plate.
• Culture : Most specific.
• PCR : Most sensitive.
• Serology : ↑ Antibody after 2 weeks.
Treatment :
• Macrolides : Azithromycin, Erythromycin.
• If allergic to macrolides : Cotrimoxazole.
Vaccine : Acellular Pertussis vaccine + Diphtheria toxoid + Tetanus toxoid.
3. Brucella 00:11:45
Brucella abortus : Brucella melitensis :

• Seen in buffalos, cows. • Seen in camels, sheep.
• Capnophilic. • M/C cause & most severe disease.
Features :
• Non motile. • Facultative intracellular.
• Non capsulated. • Urease positive.
• Strict aerobe.
Source of infection :
1. Ingestion of unpasteurised milk.
2. Inhalation of aerosols.
3. Handling infected animal products.
Clinical presentation :
Brucellosis (Malta fever/Undulent fever/Mediterranean fever/Gibraltar fever) :
• Fever, night sweats, myalgia.
• Anemia, depression.
• Focal disease :
a. Non-caseating granuloma. d. Genitourinary disease : Salphingitis/
b. Osteomyelitis. epididymitis (Infertility).
c. Arthritis. e. Meningoencephalitis.
f. Hepatitis.

Diagnosis : ----- Active space -----

1. Culture (Gold standard) : Blood/bone marrow/biopsy.
2. Serology : Standard agglutination test (Detects IgM antibodies).
3. Nucleic acid amplification test : PCR.
Treatment :
Gold standard : (Streptomycin + Doxycycline) × 6 weeks.
WHO recommendation : (Rifampicin + Doxycycline) × 6 weeks.
4. Hemophilus influenzae (Hi) 00:20:17
Features :
Non-motile.
Fastidious organism : Needs Factor X & V.
6 serotypes : A, B (Most dangerous), C, D, E, F.
Growth media :
1. Fildes agar.
2. Chocolate agar.
3. Levinthal medium.
4. Blood agar + S. aureus streak (Satellitism).
Virulence factors :
Satellitism
• Lipo-oligosaccharide.
• Pili.
• IgA protease (Also produced by S. pneumonia & Neisseria).
• Capsule (Polyribosyl ribitol phosphate).
Infection :
Invasive Non-invasive
Organism Hib (95% cases) Non typeable Hemophilus
Diseases caused • Meningitis. • Otitis Media.
• Pneumonia. • Sinusitis.
• Epiglottitis. • Acute exacerbation of
• Septic Arthritis. COPD.
• Endocarditis.
Age group 2 months to 2 years. Adults.
Rx 3rd gen Cephalosporins. Amoxyclav, Macrolides.

54 04 Microbiology

Hib vaccine : Conjugate vaccine.
Biotypes :
1. H. influenzae biotype aegypticus : Causes pink eye → Brazilian purpuric fever.
2. Hemophilus ducreyi :
• Sexually transmitted.
• Fastidious : Grows on Chocolate agar + Isovitalex and Fetal calf serum.
• IP : 4-10 days.
• C/f : Multiple painful ulcers (Chancroid) + Enlarged,
tender LN.
• Diagnosis :
a. Gram stain : School of fish appearance.
b. Culture.
c. PCR : Most sensitive. School of fish appearance
5. Mycoplasma 00:30:01
Features :
Smallest pathogenic bacteria : 0.2-0.3 microns.
Filtrable through standard bacterial filters.
Classified under Mollicutes (No cell wall → Hence resistant to antibiotics acting
on cell wall).
Jumping jokers of microbiology (Contains sterols in cell membrane).
Fastidious organism.
Culture :
Medium : PPLO medium (Pleuro Pneumonia Like Organism) + Penicillin/Thalium
acetate (Selective agent).
Colonies : Mulberry shaped/Fried egg colonies.
Note : Diene’s stain used to visualise the colonies.
Mycoplasma pneumoniae :
M/C cause of atypical pneumonia (Walking pneumonia).
Complications :
• Transient urticaria. • DIC.
• Erythema nodosum. • Meningitis.
• Steven Johnson syndrome. • Encephalitis.
• Autoimmune hemolytic anemia.


1. Detect antibodies :
• Specific : ELISA, IFA.
• Non-specific (Heterophile antibody test) :
a. Cold agglutination test.
b. Streptococcus MG test.
2. NAAT.
3. Culture : Very slow (> 1 week).
Treatment : Macrolides (DOC).
6. Legionella pneumophilia : 00:38:04
Features :
Facultative intracellular.
Non-capsulated, motile.
Natural & aquatic reservoir.
Fastidious : BCYE agar (Buffered Charcoal Yeast Extract).
No human to human transmission.
Mode of infection : Microaspiration & inhalation.
Serotypes 1, 4, 6 cause human disease (80% cases → Serotype 1).
Symptoms Findings
Variable infiltration on chest X-ray.
Respiratory symptoms.
Hyponatremia.
Mental confusion.
Hypokalemia.
Diarrhoea.
↑ Liver enzymes.
Note : Milder form → Pontiac disease.
Investigations :
1. NAAT : Most sensitive.
2. Culture : BYCE agar.
3. Urine antigen test (Highly specific for serotype 1) : ELISA/ICT
(Immunochromatography test).
Note : Urine antigen test also done in Streptococcus pneumonia and
Legionnaires disease.
Treatment : Macrolides (DOC).

56 05 Microbiology
Obligate intracellular
Bacteria : All viruses Fungi : Protista :

• T. pallidum. Pneumocystis Toxoplasma,
• Rickettsia & related genera Rhinosporidium.
(except Bartonella).
• Chlamydia.
• Tropheryma whipplei.
• M. leprae.
• Spirillum minus.
Note : Rat bite fever is caused by S. minus, Streptobacillus moniliformis.
Rickettsia & related genera 00:01:51
• Gram -ve coccobacilli; Non-motile.

• Basophilic inclusion bodies + : Stained by basic dyes like Giemsa.
• Obligate intracellular except Bartonella.
• Transmitted via arthropods except Coxiella.
• Infect endothelial cells except Anaplasma & Ehrlichia.
• C/F : Fever, rash, vasculitis.
1. Vasculitis is not seen in Anaplasma & Ehrlichia.
2. Rash : Not seen in Coxiella.
Rickettsia & related genera .................
Rickettsia Orientia Coxiella Bartonella Anaplasma & Ehrlichia
Typhus fevers. Scrub/ bush Q fever. Oroya fever. Monocytic ehrlichiosis

Spotted fevers. typhus. Peruvian wart. & granulocytic
Cat scratch disease. anaplasmosis.
Bacillary angiomatosis.
Trench fever.
Endocarditis.

Typhus fever : ----- Active space -----
R. prowazekii R. typhi
Disease Epidemic typhus/ Gaol fever. Endemic typhus.
Transmitted by Body louse- feces. Rat flea- bite/ feces.
(Pediculus humanus corporis) (Xenopsylla cheopis)
Reservoir No extra-human reservoir. Rodents.

Exception : Flying squirrels.
C/F Fever, headache, myalgia, rash starting Milder symptoms.
from trunk (doesn’t involve palms & soles).
Delirium, stupor, coma.
Latency/ Brill-Zinsser disease/ Recrudescent fever. No latency.
Reactivation
Diagnosis • Neil Mooser reaction : -ve in epidemic typhus; +ve in endemic typhus.
• Weil Felix test : Develops OX-2, OX-19 antibodies of Proteus vulgaris.
• Specific antibody test by Indirect fluorescent antibody test (IFAT).
Rx DOC : Doxycycline.
Mnemonic : Epidemics in humans are Profoundly Lousy.
Endemic disease in rodents are tough but fleeting.
(Epidemic typhus : Reservoir - Human, Agent - Prowazekii, Vector - Louse).
(Endemic typhus : Reservoir - Rodents, Agent - R. typhi & Vector - Ratflea).
Louse transmitted organisms (via feces) Flea transmitted organisms

• Bartonella quintana : Trench fever. • Bartonella henselae (via cat flea) : Cat
• Borrelia recurrentis : Relapsing fever. scratch disease, Bacillary angiomatosis.
• Rickettsia prowazekii : Epidemic typhus. • Yersinia pestis (via rat flea).
Mnemonic : TREP (TR-RE-EP) the louse. • Rickettsia typhi (via rat flea).
Spotted fevers :
R. rickettsii R. conori R. akari
Disease Rocky mountain spotted Indian tick typhus, Rickettsial pox.
fever (RMSF) Boutonneuse fever
Transmission Hard tick-bite Hard tick-bite Gamasid mite
Reservoir Ticks & rodents Ticks & rodents
C/F Rash starting from wrist & sole. Eschar/Tache Noire; Eschar;
Fever, Rash. Papulovesicular rash
Severe vasculitis :
headache, (varicelliform
Hemorrhages; gangrene,
myalgia + rickettsiosis).
encephalitis.

58 05 Microbiology
----- Active space ----- Diagnosis • Neil Mooser reaction : +ve in all.
• Weil Felix test : Develops OX-2, OX-19 antibodies of Proteus vulgaris.
It is -ve in Rickettsial pox.
• Specific antibody test by IFAT.
Note : Hemorrhagic rash in feet & soles

is also seen in syphilis, HFMD.
Gangrene seen in Eschar

epidemic typhus, RMSF
Orientia tsutsugamushi & O. chuto :

Orientia tsutsugamushi & O. chuto
(M/c rickettsial disease in India)
Disease Scrub typhus.
Transmission Chiggers (larvae) of Trombiculid mite (Transovarian transmission +).
Reservoir Rodents, birds.
C/F Eschar, Fever, truncal rash, tender lymphadenopathy, hepatosplenomegaly.
~25% : Develops meningitis/ encephalitis.
Diagnosis Weil Felix test : Develops OX-K antibodies of Proteus mirabilis.
Specific antibody test by IFAT.
PCR (from eschar scrapings).
Note : Eschar is seen in :
Spotted fevers (not RMSF), Scrub typhus, Cutaneous anthrax
and at the site of brown recluse spider bite.
Coxiella burnetii : Trombiculid mite
Coxiella burnetii
Properties • Exist as spore-like form → Survives Holder method of pasteurization.
• Shows phase variations : Phase I, II antigens.
• No rash/ role of arthropods.
Disease Q (query) fever.
Transmission Raw milk, infected animal products, aerosols, percutaneous.
Reservoir Domestic animals.
C/F • 60% : Asymptomatic.
• ~5% : Hepatitis or meningitis, encephalitis or atypical pneumonia.
• Acute Q fever in pregnancy → Spontaneous abortion.
• Culture negative endocarditis (CNE).

Diagnosis Acute : IgM titer (against phase II Ag); Blood PCR. ----- Active space -----
Chronic : IgG titer (against phase I Ag); Blood culture & PCR.
Rx Acute : Doxy x 2 wks.
CNE : Doxy + (Hydroxychloroquine/ Rifampicin/ Ciprofloxacin) x 2 yrs.
Bartonella spp. :
Not obligate intracellular; Motile.
Bartonella henselae Bartonella quintana B. bacilliformis
Trans- Cat flea feces/ cat scratch/ Feces of body louse. Bite of sandfly
mission bite. (Lutzomyia).
Reservoir Human Human Cats (zoonotic)
Disease • Typical : Cat scratch • A/k/a Trench fever or • Acute :
disease. HPE : Stellate Shin bone fever or 5 Oroya fever/
granulomas with necrosis. days fever. Carrion’s disease.
• Atypical : Parinaud’s • In native & prosthetic • If untreated :
oculoglandular syndrome. valves : CNE Peruvian wart/
• HIV : Bacillary angiomatosis • In immunodeficient : Verruga peruana.
(resemble Kaposi sarcoma). Bacillary angiomatosis.
Diagnosis • Serology (M/c) : IFAT, ELISA; Serology
• Warthin starry stain;
• PCR of pus aspirate.
Rx • Typical : Doxy/ Azithro x 5 d. Doxycycline
• Atypical : Doxy + Rifampicin.
• HIV : Doxy/ Azithro x 1-2 m.
Clumps of Bartonella on Sandfly

Warthin Starry stain
Causes of culture negative endocarditis :

• Prior antibiotics (M/c).
• Fastidious organism/ slow growers : HACEK, Bartonella, Brucella, fungus.
• Obligate intra-cellular : Coxiella, Chlamydia psittaci, Tropheryma.

60 05 Microbiology

Ehrlichia & Anaplasma :
Ehrlichia & Anaplasma
Properties Infect blood cells (not endothethial cells).
Disease Ehrlichia chaffeensis, E. muris : Human monocytic ehrlichiosis.
E. ewingii, A. phagocytophilum : Human granulocytic ehrlichiosis/ anaplasmosis.
Vector Hard tick.
C/F Fever, chills, myalgia, rash.
Diagnosis PBS : Morula (mulberry shaped inclusion bodies), leucopenia, TCP ± anemia.
PCR.
Morula in a monocyte Morula in a neutrophil

(E. chaffeensis, E. muris) (E. ewingii, A. phagocytophilum)
Chlamydia 00:37:43
• Energy parasites (no enzymes for ATP synthesis).

• Muramic acid/peptidoglycan - Absent.
• Biphasic life cycle :
1. Extracellular stage (infectious) : Elementary body (metabolically inactive).
2. Intracellular stage : Reticulate body (metabolically active).
• Has Basophilic inclusion bodies.
• Stained by : Giemsa. Iodine → Used for demonstrating inclusion bodies of
C. trachomatis (contains glycogen).
• Culture : HeLa & McCoy cell lines, Yolk sac of Hen egg.
C. psittaci C. pneumoniae
Disease Atypical pneumonia (ornithosis), Acute : Atypical pneumonia, sinusitis,
septicemia, CNE. pharyngitis.
Chronic : Atherosclerotic plaques (CAD).
Transmission Aerosols containing urine, feces, oral Aerosols.
secretions of birds.
Reservoir Birds (parrots, pigeons, turkeys). Human.
Diagnosis • Inclusion bodies : Levinthal Cole
Lillie (LCL) bodies.
• Serology (M/c).
• PCR.
Rx Doxycycline X 2-3 wks.

Chlamydia trachomatis ----- Active space -----
TRIC biovar LGV biovar
L1, L2, L3
A, B, Ba, C D-K
Lymphogranuloma
Trachoma Genital chlamydiasis venereum
D-K L1, L2, L3

Genital chlamydiasis : Lymphogranuloma venereum :
• M/c bacterial STI. • 1°: Painless papulo-vesicular genital
• M/c cause of urethritis, ophthalmia lesion.
neonatorum. • 2°: Painful enlarged LN (groove sign).
• Males : Urethritis (watery discharge), • 3°: Lymphatic obstruction (genital
epididymitis, proctitis. elephantiasis - K/a Esthiomene in
• Females : Cervicitis, endometritis, infertility, females).
Fitz-Hugh Curtis syndrome, PID, abortion.
• Swimming pool conjunctivitis, infant pneumonia.
Diagnosis : NAAT. Diagnosis : Miyagawa’s granulocorpuscles
(elementary bodies), Serology, NAAT.
DOC : Doxycycline. Rx : Doxy/ Erythromycin x 3 wks.
In neonates : Erythromycin.
Causes of urethritis :
• Gonococcal urethritis (purulent discharge) : Gonococcus.
• Non-gonococcal urethritis : C. trachomatis D to K (M/c), Ureaplasma,
Mycoplasma genitalium.
Spirochetes 00:51:20
• Gram -ve slender spirals.

• Endoflagellae +.
• Family Spirochetaceae : Treponema, Borrelia.
Family Leptospiraceae : Leptospira.

62 05 Microbiology

Leptospira spp. :
Leptospira spp.
Properties • Seen under dark ground microscope (DGM).
• Closely wound spiral with 1 or both ends hooked (Umbrella handle/
shepherd crook appearance).
• Culture medium : EMJH, Korthoff, Fletcher’s, Stewart’s → Forms Dinger’s ring.
IP 2-20 days.
Transmission Urine contaminated water.
Occupational infection (Rice field farmers, sewer workers).
Reservoir Rodents; domestic animals.
Disease • 40% : Asymptomatic.
• 60% : Mild febrile illness.
5-10% : Weil’s disease/ ictero hemorrhagic fever/ hepatorenal syndrome.
2 stages of 1. Septicemic stage (5-7 days) : Fever, chills, myalgia, conjunctival suffusion,
disease calf tenderness, pretibial rash.
2. Immune stage : Jaundice, hemorrhages, renal dysfunction, meningitis.
Diagnosis • Blood : +ve in 1st week.
• Urine, CSF : +ve in 2nd week.
• DGM, PCR, culture.
• Serology : Microscopic agglutination test/ MAT (gold standard).
Rx Anicteric febrile illness : Doxy/ Amoxicillin/ Ampicillin orally.
Weil’s disease : IV Penicillin/ Ceftriaxone.
EM of leptospira DGM of leptospira

Treponema pallidum : ----- Active space -----
Treponema pallidum
Properties • Seen under DGM.
• Corkscrew motility.
• Grow on rabbit testis.
IP 9-90 days.
Disease Syphilis.
Stages of • Primary : Painless, indurated genital ulcer + B/L non-tender rubbery LNAP.
disease • Secondary (Great imitator) : Symmetric rash (palms & soles), mucous
patches, generalized LNAP, condyloma lata, meningitis, hepatitis.
• Latent :
Early latent (<1 yr after infection) : Relapses of manifestations of 2° stage.
Late latent : >1 yr after infection.
• Tertiary : Gummas, aortitis, AR, aotic aneurysm (ascending aorta).
Neurosyphilis : Tabes dorsalis, General paresis of insane.
Diagnosis • 1° & 2° : DGM, silver stains, DFAT, PCR, serology.
Serology : 2 step approach.
1. Screening test : Non- treponemal test → VDRL/ RPR test.
2. If +ve, Confirmatory test : Treponemal tests → TPHA/ TPPA/
FTA-ABS/ TPI.
3. Follow up : VDRL/RPR → Look for 4 fold fall in Ab titers after 6 months.
• Neurosyphilis : VDRL (more specific) > FTA-ABS.
Rx Early syphilis (1°, 2°, early latent) : Benzathine Penicillin IM single dose.
Late syphilis : Benzathine Penicillin IM x 3 wks (1 dose/ wk).
Neurosyphilis : Aqueous crystalline penicillin x 14 days.
Pregnancy : Benzathine Penicillin.
Note :
Most specific test : TPPA.
Most sensitive test : FTA-ABS
Causes of biological false positives :

• Acute : Pregnancy, infectious mononucleosis, measles, malaria, chicken pox,
viral hepatitis, recent immunizations, acute HIV infection.
• Chronic : SLE, Lepromatous leprosy, Collagen disorders, Narcotic addictions,
Malignancies like myeloma.
Reverse testing of syphilis : Automated treponemal test is done first followed by

confirmatory non-treponemal test.

64 05 Microbiology

Borrelia :
• Borrelia (Broader) : Seen under light microscope.
• Relapsing fever (louse borne & tick borne), Lyme’s disease.
Louse borne Tick borne
Lyme’s disease
Relapsing fever (LBRF) Relapsing fever (TBRF)
Caused B. recurrentis B. duttoni B. burgdorferi.
by (Antigenic variations) (Antigenic variation)
Trans- Louse (feces). Soft tick (bite). Hard tick (bite) :
mission Ixodid ricinus complex.
Reservoir Humans. Rodents, birds, squirrels Rodents; deer
No animal reservoir.
Aka epidemic RF. Aka endemic RF. • 1°: Erythema migrans.
• 2° (Latest great imitator) :
Lymphadenopathy, meningitis,
fluctuating AV blocks.
• HSM, meningitis, LNAP, hemorrhages (more • 3° (Stage of persistence) :
severe in LBRF) Arthritis, encephalopathy,
• Remitting RF (more severe in TBRF). 7th nerve palsy.
Diagnosis Blood smear (collected during febrile period). Serology : 2-step approach.
Culture : Barbour Stoenner Kelly (BSK) media Screening by ELISA.
(Most specific). Confirmation by Western blot.
PCR : Most sensitive.
Rx Tetracycline/ Doxy Tetracycline/ Doxy x • No CNS/CVS involved :
once. 10 days. Doxy (2 wks - 2 Months).
• CNS/CVS involved :
i.v. Penicillin/Ceftriaxone.
Dorsal shield
Soft tick Hard tick Wright-Giemsa stained peripheral

blood smear of a patient with TBRF
Erythema migrans/ Bull’s eye rash

Campylobacteriaceae 01:15:40 ----- Active space -----
• Gram -ve spirals.

• Microaerophilic.
• Comprises of Campylobacter & Helicobacter.
• Catalase & oxidase +ve.
• Culture medium : Skirrows, Butzler medium, Campy BAP.
Campylobacter spp. Helicobacter pylori

Properties • M/c species : C. jejuni (optimum • Lophotrichous.
temp of growth 42°C). • Maximum urease producer.
• Amphitrichous; Darting motility. • Normal habitat : Gastric antrum.
• IP : 2-7 days. • Virulent factors : cag PAI, vacA,
urease.
• Protective in GERD, Barrett’s
esophagus, esophageal adenoCA.
Transmission Poorly cooked Poultry, raw milk, Feco-oral.
feces contaminated food & water.
C/F Invasive diarrhea. Duodenal > Gastric ulcer.
Complication : Guillain Barre Syndrome. Gastric adenocarcinoma, MALTomas.
Diagnosis • Gram stain (stool) : Gull wing &
‘S' shaped forms.
• Culture : Gold standard.
Rx DOC : Azithromycin.
Electron microscopy Warthin starry stain of

of H. pylori gastric Bx showing H. pylori

66 06 Microbiology
Immunity 00:00:33
Innate immunity Adaptive /acquired immunity

No lag phase. Lag phase (4-7 days to generate response).
Non specific. Specific.
No memory response. Memory response.
• Diversity of receptors limited. • Unlimited diversity.
• Pattern recognition receptors : • T cell & B cell receptors.
Recognize pathogen associated
molecular patterns (PAMP).
Innate immunity
Physical barriers Chemicals Commensal flora Immune cells :

• Monocytes.
Complement Antimicrobial peptides Acidic • Macrophages.
proteins in blood & secretions pH of • Mast cells.
stomach • Nk & NKT cells.
• Granulocytes.
• γδ T cells.
• B1 & MZ B cells.
Adaptive immunity
Humoral Cell mediated

B2 cells αβ T cells
Adaptive immunity
Active Passive
Natural Artificial Natural Artificial

(Follows infection) (Vaccination) (Mother to child (Preformed
transfer) immunoglobulins)

Lymphoid organs : ----- Active space -----
Primary Secondary
Bone marrow. Lymph nodes.
Thymus. MALT.
Spleen.
Lymph node Spleen

T cell dependent area Paracortex. Periarteriolar lymphoid sheath (PALS).
B cell dependent area Cortex. Follicles around PALS.
Determinants of immunogenicity/antigenicity :
1. Chemical nature : Proteins > Polysaccharides > Nucleic acids > Lipids.
2. Molecular weight : 5000-10,000 daltons for detection.
3. Foreignness.
4. Susceptibilty to host enzymes.
Classes of antigens :
T independent T dependent
B cells secrete antibody without T helper cell signalling for antibody
T helper cell signal. production.
Antigens : Carbs, lipids, nucleic
Antigen : Complex proteins.
acid, simple proteins.
No class switching. Class switching +.
No affinity maturation. Affinity maturation +.
No memory response. Memory response +.
Superantigens :
• No processing by antigen presenting cells (APC).
• Binding : Directly to lateral part of MHC- II. MHC
• Recognition : V-β part of T-cell receptor.
Super
• Activates : 5-10 % of TH cells & initiates cytokine storm. Ag
• Example :
a. Toxic shock syndrome toxin.
b. Staph enterotoxin A, B.
c. Pyrogenic/erythrogenic streptococcal toxin.

68 06 Microbiology

Heterophile antigens :
Test Disease Antibodies against
Paul Bunnel test. Sheep RBC.
Infectious mononucleosis.
Monospot test. Horse RBC.
Cold agglutination. Group O RBC.
Mycoplasma pneumoniae.
Streptococcus MG test. Group F strep.
Weil felix test. Rickettsia infection. O antigen of proteus.
Antibodies 00:17:58
Classes of heavy chains : a, g, m, e, d.

Classes of light chains : l, k.
Antigen binding part : Variable part of heavy & light chain (VH & VL).
Class determining part : Constant region of heavy chain (CH).
Variable
part
Variable part :
3 hypervariable loops each in VH & VL.
Complementarity-determining region (CDR)/
Paratope. Constant
part
Antibody structure
Action of papain on antibody :

Breaks antibody above hinge region.
2 Fab + 1 Fc derived.
Action of pepsin on antibody : Action of Papain

Breaks antibody below hinge region.
1 F(ab1)2 + peptides.
Action of Pepsin

Heavy Crosses Complement ----- Active space -----

Antibody Valency Special characters
chain placenta fixation
IgM
pentamer m
linked Yes Largest
10 No
by J (Activates (9,00,000 Dalton).
chain. classical
complement
pathway). • Fc part binds to
g
IgG 2 Yes phagocytes.
• Most abundant (80%).
IgA • Dimer → Mucosal
dimer a secretions.
J 4 Monomer → Serum.
chain. • Activates alternate
complement pathway.
No No
• Mediates Type 1 HSR
(Mast cells, basophils).
IgE e 2
• Least abundant
(0.002%).
IgD d 2 Acts on B cell receptors.
Functions of antibody :
1. Opsonization : IgG (Receptor is Fc of IgG).
2. Antibody dependent cell cytotoxicity : IgG.
3. Neutralization : IgG & IgA.
4. Activation of classical complement pathway : IgM > IgG (IgG3 > G1 > G2).
Complement system 00:30:42
Complement system
Classical Lectin Alternate

(Adaptive immunity) (Innate immunity)
Note : C1qr2s2→ C1 complex.

70 06 Microbiology
----- Active space ----- Classical pathway Lectin pathway Alternate pathway
Activated Antigen binding Mannose binding lectin Lipopolysaccharide,
by to Fc portion of binding to mannose on Viral envelopes,
antibody. antigen. Fungal wall.
Lectin Classical Alternate
Membrane Attack
Complex (MAC)
Complement pathways
Classical/lectin pathway Alternate pathway

C3 convertase C4b2a. C3bBb (C3b + Factor B, D).
(Amplification)
C5 convertase C4b2a3b. C3bBb3b.
Complement proteins :
5% of serum proteins.
Most abundant complement : C3.
Functions :
1. Membrane attack complex : C5-9. 5. Regulatory proteins :
2. Chemotactic : C5a & C3a. • Decay Accelerating Factor (DAF),
3. Opsonization : C3b. membrane cofactor protein, CR-1 :
4. Anaphylotoxin : C5a > C3a > C4a. Prevents binding of C3b to normal
host cells.
• Factor I & H : Breaks down C3b.
• C1 esterase : Breaks down C1
complex.
Deficiency : ----- Active space -----
Deficient Disease
C1 esterase inhibitor. Hereditary angioneurotic edema.
C1, C2, C4. SLE like disease.
C3. Recurring pyogenic infections.
C5-9. ↑ susceptibility to neisseria.
CD59, Decay accelerating factor. Paroxysmal nocturnal hemoglobinuria.
Antigen-antibody reactions 00:42:15
Specific.
No denaturation.
Reversible.
Bonds formed : Ionic bonds, van der Waals forces, hydrophobic interactions &
hydrogen bonds.
Process Examples
Precipitation Soluble antigen + VDRL : Slide flocculation.
reactions Specific antibody Kahn : Tube flocculation.
Precipitation in gel : Immunodiffusion.
Types :
• Oudin : Single diffusion one dimension.
• Oakley fulthorpe : Double diffusion one
dimension.
• Mancini : Radial immunodiffusion.
• Ouchterlony : Double diffusion two
dimensions.
Agglutination Particulate antigen Blood grouping.
reactions + Specific antibody Heterophile Agglutination Tests (HAT).
Widal test, Standard Agglutination Test
(SAT).

72 06 Microbiology

Marrack’s lattice hypothesis :
Prozone Postzone
Process Examples
Complement Antigen + antibody + complement
Wasserman test.
fixation fixation.
Toxin neutralization → Eg. : Nagler reaction.
Neutralization
Viral neutralization → Eg. : Plaque reduction neutralization
reactions
test.
ELISA Enzyme labelled antibody
-
CLIA Substrate of enzyme emits light in CLIA.
Fluorescent • Direct FAT.
Fluorochrome labelled antibody.
antibody test (FAT) • Indirect FAT.
Types of ELISA :
Process Uses
Substrate
Enzyme conjugated to antibody
Direct Antigen detection.
directed towards the antigen.
Enzyme conjugated to secondary Antigen & antibody

Indirect
detection antibody. detection.
Antigen trapped b/w capture &

Sandwich Antigen detection.
detection antibody.
2 antigens compete with each

Competitive -
other for antibody.

Fluorescent antibody tests : ----- Active space -----
Western blot :
To detect proteins in specimen.
1. Antigens separated by gel electrophoresis.
2. Transferred to nitrocellulose.
3. Primary antibody added.
4. Membrane washed & secondary conjugated antibody added.
5. Substrate added & colour change detected.
Immunochromatographic test/lateral flow assay :

Rapid tests.
Eg. : Malaria, SARS-COV-2, HBs Ag detection.
MHC antigens 00:52:38
Encoded by MHC locus.

Location : Short arm of chromosome 6 (6p).
Function : Presents peptide to T cells.
Location Encodes
HLA-I Telomeric end of 6p. MHC I antigens.
HLA-II Centromeric end of 6p. MHC II antigens.
C2 & C4, TNF-α, Heat shock

HLA-III B/w HLA-I & II.
proteins, Factor B.

74 06 Microbiology
----- Active space ----- MHC-I antigens MHC-II antigens

Encoded by HLA A, B & C. HLA-DP, DQ, DR.
Structure
b2 microglobulin
All nucleated cells &

Expression on Antigen presenting cells (APC).
platelets.
Assembly with
Endoplasmic reticulum. Endosome.
peptide
Cytosolic/endogenously
Peptide Peptides of exogenous structure.
synthesised peptide.
Presents peptides to Presents peptides of exogenous
Function
cytotoxic T cells. antigen to T helper cells.
Peptide binding
α-1 & α-2. α-1 & β-1.
cleft
Antigen presenting cells :

Signals for activation of T helper cells :
• 1st signal : Peptide recognition in groove of MHC -II.
• 2nd signal : CD-28 of T helper cell binding to B7 (CD 80/81) molecule of APC.
Antigen
MHC-II T cell receptor
Antigen presenting
cell Naïve TH
CD4
CD80/81 CD28
(B7) Activated TH

APC ----- Active space -----
Professional Non professional

Dendritic cells (Most efficient). Fibroblasts, glial cells.
B cells. Vascular endothelial cells.
Macrophages. Pancreatic β-cells.
Thymic & thyroid epithelial cells.
Natural Killer cells/Null cells :

Part of innate immunity.
Markers : CD16 (FcR for IgG) , CD56.
Function : Defence against virus infected & cancerous cells.
Inhibitory receptors on Nk cells bind to MHC-I.
Show antibody dependent cell cytotoxicity (Release Perforins & Granzymes →
Apoptosis).
Cytokines secreted : IFN-γ, TNF-α.
Cytokines activating Nk cells : IL-2, IL-12, IL-15, IFN-α & β
T cells 01:04:49
Originate : Bone marrow.

Maturation : Cortex & medulla of thymus.
Marker : CD3 (Pan T cell marker).
Form E rosettes with sheep RBC.
Types :
1. αβ (90-95 %) :
• Part of adaptive immunity.
• 2 subtypes :
a. Cytotoxic T cells : Marker CD8.
b. Helper T cells : Marker CD4.
2. γδ (1-10%) : Part of innate immunity, recognise lipids & present in mucosa.
Diversity of T cell receptors : Somatic rearrangements/gene recombination.

76 06 Microbiology

Positive selection :
Occurs in thymic cortex.
T cells which recognize either of MHC allowed to mature further in thymus &
rest undergo apoptosis.
Negative selection :
Occurs in thymic medulla.
T cells which do not recognise self antigens receive survival signal.
Responsible for self tolerance.
Autoimmune regulator (AIRE) : Mutation of AIRE results in autoimmunity.
T cell Function Secrete

Th1
Activates macrophages. IL-2, IFN-γ.
Activates B cells.
Th2 IL-4, IL-5 & IL-13.
Th 17 Activates neutrophils.
IL-17, IL-21 & IL-22.
T reg Regulatory role.

Induce apoptosis of cells with intracel-
Cytotoxic T cells Activates caspases.
lular pathogens.
Innate immunity
NK T cells
Has αβ T cell receptor.
B cells 01:16:14
Maturation : Bone marrow.

Markers : CD19 (Pan B cell marker).
Form EAC rosettes.
Not MHC restricted.
B cell receptor (BCR) :

Diversity of B cell receptor : Occurs by somatic rearrangements/gene
recombination.
2 classes : IgM & IgD → Occurs due to alternate RNA splicing.

Negative selection : By AIRE. ----- Active space -----
Isotype switching :
B cells secrete IgM.
Signal from follicular helper T cells (CD40 binding to CD40L) → B cells secrete
other classes of antibodies → Isotype switching/class switching.
CD40L absence : Hyper IgM syndrome.
Affinity maturation :
Low affinity antibodies→ High affinity antibodies.
Cause : Somatic hypermutation in secondary folliclles.
Mediated by follicular dendritic cells.

78 07 Microbiology
Virology 00:00:11
Basic structure of a virus :
Capsid :
• Protein coat → made of capsomeres.
• Arrangement of capsomeres determine
symmetry of the virus.
DNA Viruses.
ds DNA virus ss DNA virus
Enveloped Non-enveloped Icosahedral

Non-enveloped
Complex Icosahedral Icosahedral
Nuclear replication.
cytoplasmic nuclear replication nuclear replication
replication Parvovirus
• Hepadna virus • Papilloma virus
Poxvirus • Herpes virus • Adenovirus.
• Polyoma virus.
RNA Viruses.
ss RNA virus ds RNA virus
Positive sense Negative sense Icosahedral

Non enveloped non-enveloped
Enveloped Enveloped Enveloped
icosahedral helical icosahedral Reovirus
Icosahedral Helical • Arena
Delta
• Picorna • Toga Corona • Bunya
• Astro • Flavi • Paramyxo
• Calici • Matona • Orthomyxo
• Hepe • Retro • Filo
• Rhabdo
All RNA viruses multiply in cytoplasm, except : ----- Active space -----
1. Measles.
2. Orthomyxovirus. Replication partly in cytoplasm
3. Retrovirus. & partly in nucleus.
• Smallest pathogenic virus : Parvovirus B19.

• Largest pathogenic virus : Pox virus.
• Smallest genome virus : HDV.
• Smallest DNA genome virus : HBV.
• Largest genome virus : Pox virus.
• RNA virus with the largest genome : Coronavirus.
• Diploid genome virus : Retrovirus.
• RNA virus which replicates through DNA intermediate : Retrovirus.
• DNA virus which replicates through RNA intermediate : Hepadnavirus.
Note :
• Both Retrovirus and Hepadenavirus have reverse transcriptase.
• Smallest genome virus with independent replication : HBV.
Virus with segmented genome can undergo genetic reassortment :

• Reo virus (10-12).
• Orthomyxo virus (7-8).
• Bunya virus (3).
• Arena virus (2).
Genetic reassortment : If 2 strains of a virus co-infect the same host cell →

new strains can arise during the assembly.
Filovirus : Ebola, Marburg. Wheel like : Rotavirus. Brick shaped : Pox.

80 07 Microbiology

Inclusion bodies :
Inclusion bodies Viruses

Guarnieri (C). Variola virus.
Henderson Patterson bodies (C). Molluscum contagiosum.
Cowdry A (N). Herpesvirus.
Cowdry B (N) Adenovirus.
Negri (C). Rabies (Maximally seen in hippocampus).
Torres (N). Yellow fever virus.
Owl eye (N). Cytomegalovirus (CMV).
Note : Both intranuclear and intracytoplasmic inclusion bodies are seen in CMV and measles
infection.
(C)→ Cytoplasmic, (N)→ Nuclear.
Negri bodies Owl eye inclusion body Warthin Finkeldey giant cells
Tzank smear : Multinucleated giant cells (HSV/VZV).

Vector borne viruses :
Rodent borne viruses : Hard tick borne Aedes borne viruses : Culex borne virus :
viruses :
• Arenavirus : Lassa • KFD (India). • • JE virus (Flavi)
Chikungunya (Toga).
fever virus. • TBE. • • West Nile virus
Dengue (Flavi).
• Bunya : Hantavirus. • CCHF (India). • Yellow fever (Flavi).
(Flavi).
• • St. Louis
Zika (Flavi).
encephalitis
(Flavi).
TBE → Tick borne encephalitis : Russian spring summer encephalitis (RSSE).
Viruses causing hemorrhagic fever :

• Dengue (Flavi). • Hanta (Bunya).
• Yellow fever (Flavi). • South American HF (Arena).
• KFD (Flavi). • Ebola (Filo) → most fatal.
• CCHF (Bunya). • Marburg (Filo).
DNA viruses 00:17:50 ----- Active space -----
Parvovirus B 19 :
Properties :
Receptor : P antigen on erythroid precursors.
Clinical features :
Mostly asymptomatic.
When symptomatic (D/t 10 Parvovirus infection) :
• Young children : 5th disease/erythema infectiosum.
• Adolescent : Papulopurpuric gloves & socks syndrome.
• Adult : A/c, symmetric small joint arthropathy.
• Hemolytic anemia : Aplastic crisis.
• Immunodeficiency : Pure red cell aplasia. Slapped cheek appearance
• Pregnancy : Non immune fetal hydrops.
• Hemophagocytic syndrome.
Papilloma Virus : 00:20:56
Properties :
Circular ds DNA.
Oncogenic.
High risk (HPV 16,18) & low risk (HPV 6,11).
Structural proteins : L1 (Incorporated in vaccine) & L2.
Oncoproteins :
• E6 → inactivates product of p53.
• E7 → inactivates product of RB.
Clinical features :
Mostly asymptomatic.
Symptomatic
Skin warts/ Genital warts Respiratory Epidermodysplasia Anogenital &

verrucae (6, 11). papillomatosis verruciformis oropharyngeal
(6, 11). (5, 8). carcinomas
(16, 18, 31, 33, 45,
52, 58).

82 07 Microbiology

Viral Hepatitis : 00:24:26
HBV :
Dane particle.
Properties :
Enveloped, partial ds DNA.
Replicates through an RNA
intermediate : Pregenomic RNA.
M/c cause of hepatocellular

carcinoma & end stage liver dis-
ease worldwide.
Structure :
Capsid protein : HBcAg.
Envelope has HBsAg (Australia
antigen).
Genome has 4 genes :
1. S gene : HbsAg.
2. P gene : DNA polymerase.
3. C gene : Encodes HBcAg & HBeAg (Soluble precore protein).
4. X gene : Encodes transactivator protein (Role in carcinogenesis).
Route of transmission :
Parenteral, mucosal exposure to blood, semen, saliva, tears & cervical secretions.
Risk of transmission by needle stick exposure : 30-40%.
HBV serum markers :

Antigens
HBsAg HBeAg HBcAg

First to appear. Detectable at high Not detected in serum.
Disappears on resolution. viral replication.
Antibodies
IgM anti HBc IgG anti HBc Anti HBs Anti HBc
1 antibody to
st
Replaces IgM & Appears on
appear in serum. persists lifelong. resolution.

Interpretations :
Serum markers Inference

HBsAg +, HBeAg +, IgM anti HBc +. A/c infection.
HBsAg +, IgG anti HBc +, HBeAg +. C/c infection with high infectivity.
HBsAg +, IgG anti HBc +, HBeAg -. C/c infection with low infectivity.
Anti HBs +, anti HBc +. Immune due to previous infection.
Anti HBs +, anti HBc -. Immune due to vaccine.
Anti HBs -, anti HBc -. Susceptible to Hep B infection.
Precore mutant : Mutation in precore region of C gene : HBeAg -ve hepatitis.

Escape mutant : Mutation in ‘a’ determinant of HBsAg → not neutralised by
vaccine induced anti Hbs.
Vaccine : Recombinant HBsAg.

Protective level of anti HBs : >10 IU/mL.
HDV :
Properties :
Defective virus (Lacks HBsAg encoding gene).
Only replicates in the setting of a HBV infection :
Coinfection Superinfection
Less common. More common.
Chronicity likelihood <5%. Chronicity likelihood ~90%.
IgM anti HBc (+). IgG anti HBc (+)

84 07 Microbiology

HAV & HEV :
Features HAV HEV

Family Picornaviridae. Hepeviridae.
Aka Enterovirus 72. Epidemic non A, non B hepatitis virus.
Transmission Faecooral transmission.
IP 15-50 days.
Diagnosis IgM in serum.
Complications - Fulminant hepatitis in pregnancy.
Vaccine Formalin inactivated -
vaccine.
Herpes virus : 00:36:19

3 subtypes : HHV 8 causes :
1. α : HSV 1, HSV 2, VZV. • Kaposi sarcoma.
2. β : CMV, HHV 6, HHV 7. • Primary effusion lymphoma (PEL).
3. γ : EBV, HHV 8. • Multicentric Castleman disease.
Note : HHV 6 & 7 cause exanthema subitum/roseola infantum/6th disease.
HSV 1 & HSV 2 :

Transmitted by skin/mucous
membrane breaks.
Asymptomatic → latent in sensory ganglia.
Symptomatic : Gingivostomatitis
• Mucocutaneous lesions : Gingivostomatitis,
herpes labialis, keratitis, oesophagitis.
• Genital herpes.
• Herpetic whitlow.
• Neurological : Herpetic whitlow Herpes labialis
a. M/c cause of sporadic encephalitis worldwide : HSV 1.
b. Localized to the temporal lobe.
c. Diagnosis : CSF → PCR.
d. Rx : Acyclovir.

CMV/HHV 5/salivary gland virus : ----- Active space -----

Transmitted by close contact, sexual, blood, blood products.
Becomes latent in secretory glands & kidneys.
Cells infected with CMV : Enlarged, owl eye inclusion bodies.
Clinical features :
The infection can either be a primary CMV infection or a reactivation.
1. Infectious mononucleosis (IM) like syndrome :

Diagnosis : Detection of anti CMV IgM by
• Nucleic acid amplification test (NAAT).
• Direct fluorescent antibody test (DFAT).
2. In pregnancy :
Primary infection >> reactivation.
Risk of transplacental transmission.
Transplacental transmission : Throughout pregnancy (maximum in first half).
Cytomegalic inclusion disease :
• Microcephaly. • Deafness.
• Periventricular calcification. • Mental retardation.
• Chorioretinitis. • Petechial rashes.
Diagnosis :
a. Antenatal : Quantitative PCR of amniotic fluid.
b. NAAT (Urine, saliva, blood).
c. Anti CMV IgM.
3. In AIDS : Reactivation → Chorioretinitis (M/c), Colitis, Encephalitis.
4. In transplant recipient :
Reactivation → Fever with Leucopenia. Also cause pneumonitis, hepatitis.
Maximum chance in the first 4 months post transplant.
EBV/HHV 4 :
Transmitted by saliva.
Infects B cells → polyclonal activation → heterophile
antibodies. Atypical
lymphocytes

86 07 Microbiology

Receptor : CD 21 Aka Complement receptor 2 (CR2).
Atypical lymphocytes : Activated cytotoxic T cells.
Clinical manifestations :
• Infectious mononucleosis/ Kissings disease/ Glandular fever :
Diagnosis :
a. Paul Bunnell test : Detects antibody against sheep RBCs.
b. Monospot test : Detects antibody against horse RBCs.
Admistering ampicillin in Infectious mononucleosis patients sometimes can
produce rash.
• X linked lymphoproliferative disease/Duncan’s disease/Purtilo syndrome
Hemophagocytosis.
• Hemophagocytic lymphohistiocytosis.
• Oral hairy leukoplakia.
• Carcinomas.
Carcinomas associated with EBV :

• Hodgkin’s lymphoma. • Post transplant
• Burkitt’s lymphoma. lymphoproliferative disease.
• Primary CNS lymphoma. • NK cell/T cell lymphoma.
• Nasopharyngeal carcinoma. • PEL.
VZV/HHV 3 :
Transmitted by droplet nuclei (Airborne precautions).
Chicken pox :
Incubation period (IP) : 7-23 days.
Period of infectivity : 2 days before rash
to 5 days after onset.
Pleomorphic rash. Fig : Small pox Fig : Chicken pox
Complications :
• 20 bacterial infection (M/c).
• Ataxia.
• Pneumonia.
Rx : Acyclovir.
Vaccine : Varivax (Oka strain).
(Live vaccine). Fig : Centripetal rash. Fig : Dew drop
on rose petal.
Zoster : ----- Active space -----

Reactivation of latent VZV in dorsal root ganglion.
Clinical features : Dermatomal distribution of painful
vesicles.
M/c dermatome : D3 to L2.
Complications :
• Post herpetic neuralgia. Fig : Lesions in herpes zoster
• Herpes zoster oticus.
• Herpes zoster ophthalmicus.
• Zoster sine herpete (pain in dermatome but
painless vesicles).
Vaccine :
Fig : Herpes zoster ophthalmicus
• Shingrix : Recombinant.
• Zostavax : Live attenuated.
RNA viruses 00:51:44
Flaviviridae
Hepacivirus Flavivirus
HCV Hard tick

Mosquito
Culex : Aedes : • KFD

• JE virus • Dengue • TBE (RSSE,
• West Nile virus • Yellow fever Powassan virus)
• Zika

88 07 Microbiology

JE virus vs West Nile virus :
Features JE virus West Nile virus

Transmit- Culex. Culex
ted by Blood, sexual, solid organ transplant,
breast feeding.
Reservoir • Natural : Aquatic birds/ardeid birds. Crows, jays, sparrows.
• Amplifier : Pigs. Dead end hosts : Humans & horses.
• Dead end hosts : Humans & horses.
IP 5-15 days. 5-15 days.
Clinical < 1% : Encephalitis. Fever, myalgia.
features Mortality : 25-50%. < 1% : Encephalitis, meningitis, acute
m/c cause of epidemic encephalitis in flaccid paralysis.
Asia. Mortality : 10%.
Diagnosis IgM ELISA. IgM ELISA → PRNT.
Confirmatory : Plaque reduction neu- NAAT of CSF & serum.
tralisation test (PRNT).
Vaccine Killed & recombinant vaccines. No vaccine.
Strains used :
• SA-14-14-2.
• Beijing 1, Nakayama.
Yellow fever vs KFD :
Features Yellow fever KFD

Prevalence Africa, South America. South India.
Vector Aedes. Hard tick (Haemaphysalis
spinigera).
Reservoir Primates. Rodents.
Amplifying host : Monkeys.
Clinical Fever, myalgia, jaundice, hemorrhage, Fever, myalgia, hemorrhagic
features A/c tubular necrosis manifestation,
Torres bodies (+). meningoencephalitis.
Mortality : 20%.
Diagnosis ELISA → PRNT. ELISA, RTPCR.
RTPCR on blood.
Vaccine 17 D vaccine (lifelong immunity). Killed vaccine.

Zika virus vs Dengue : ----- Active space -----
Features Zika Dengue

Transmitted by Aedes, blood, sexual, transplacental Aedes.
(Congenital zika syndrome).
Reservoir Primates. Primates.
Clinical • Fever. • Fever.
features • Rash. • Retroorbital pain.
• Headache. • Joint pain.
• Joint pain.
Complications GBS, congenital zika syndrome Severe dengue :
(Microcephaly). • Age < 12 yrs.
• DEN 1 followed by DEN 2.
• Malnourishment is protective.
Diagnosis • ELISA → PRNT. • ELISA → PRNT.
• RTPCR. • NS1 antigen.
• RTPCR.
Vaccine - Dengvaxia : Recombinant.
Paramyxoviridae : 01:04:02
1. Morbillivirus :
• Measles.
• Rubeola.
2. Rubulavirus :
• Mumbs.
• Parainfluenza 1,3.
3. Respirovirus :
• Parainfluenza 2,4.
4. Pneumovirus :
• RSV : Bronchiolitis.
5. Henipavirus (Zoonosis) : Fig : Life cycle of Nipah virus
• Nipah virus : Encephalitis & Mortality in 45-75% cases.
• Hendra virus.
Note : Parainfluenza virus causes croup.

90 07 Microbiology

Orthomyxoviridae : 01:07:26
Influenza virus :
Properties :
Hemagglutinin :
• Binds to sialic acid receptors.
Neuraminidase :
• Destroys sialic acid receptor.
Influenza A,C : has 8 segments of genome.
Influenza B : has 7 segments of genome.
Antigenic shift Antigenic drift

Due to genetic reassortment. Due to mutations.
Occurs over 10-12 years. Occurs in 1-2 years.
Sudden change in antigenicity. Gradual change.
Transmitted by : Droplet.
Complications :
• Pneumonia (M/c).
• Reye’s syndrome (Influenza B).
Diagnosis :
• Specimen : Nasopharyngeal secretions.
• Tests : RTPCR, DFAT, hemagglutination test.
Rhabdoviridae : 01:10:39
Properties :
Bullet shaped.
Envelope has glycoproteins which binds to
acetylcholine receptors.
Rabies :
Lysavirus type 1.
Transmitted by bite of rabid dog or other animals, licks on abraded skin & intact
mucosa, aerosol inhalation, organ transplantation, corneal transplantation.
IP : 1-2 months.

Clinical features : ----- Active space -----

• Furious (80%) : Hydrophobia, aerophobia, hallucinations, hypersalivation.
• Paralytic (20%) : Ascending flaccid paralysis.
• 100% mortality.
Diagnosis
Antemortem Postmortem
Sample Skin biopsy (Nape of neck), Brain biopsy, salivary
corneal smears, saliva. glands.
Tests NAAT, DFAT, culture. NAAT, DFAT, negri bodies.
Prophylaxis : Cell line derived vaccine.
Filoviridae : 01:13:46
Reported mainly from Africa. Never from India.
Ebola virus :
6 species :
• Most fatal : Zaire.
Reservoir : Bats.
IP : 2-21 days.
Clinical features : Hemorrhagic fever, DIC.
Mortality : 40-70%.
Rx : Mono/trivalent monoclonal antibodies (for Zaire).
Vaccine : Ervebo (Recombinant).
Other filovirus :
• Marburg. Reservoir : Bats.
• Ravn.
Crimean Congo hemorrhagic fever (CCHF) : 01:16:20
Belongs to Bunya virus family.

Vector : Hard tick.
Human to human transmission (+).
Amplifying host : Cattle, goat, sheep.
Geographical distribution in India : Rajasthan and Gujarat.

92 08 Microbiology
Protozoa 00:00:10
Unicellular eukaryotic organisms (Kingdom Protista).

Classified based on organs of locomotion :
Kingdom Protista
Motile Non-motile
a. Toxoplasma
Pseudopods Flagellates Ciliates
b. Babesia (Less
a. Entamoeba a. Giardia Balantidium coli common in India).
b. Free-living b. Trichomonas c. Plasmodium
amoeba. c. Leishmania d. Cyclospora
d. Trypanosoma e. Cryptosporidium
(Not seen in India). f. Cystoisospora
Pseudopods 00:02:17
Entamoeba histolytica :
Host : One (Human).
Habitat : Large intestine & caecum.
Mode of infection (MOI) : Ingestion of quadrinucleate cyst (10-100 cyst)
Excystation to form trophozoites.
Disease :
1. Asymptomatic (In 80-90%).
2. Amoebic dysentery (10-20% trophozoite invasion)
• C/F :
a. Fever.
b. Diarrhoea/dysentery.
c. Flask-shaped (Waterbottle/undermined) ulcers.
• Diagnosis :
a. Stool PCR (Gold standard).
b. Antigen (ELISA).
c. Microscopy for trophozoites & cysts.
d. Culture (Diamonds media/Krieg’s media).

3. Amoebic liver abscess : ----- Active space -----
• Male>> female : 10 : 1.
• M/C in posterior-superior aspect of right lobe of liver.
• C/F :
a. Fever.
b. Pain.
c. Heaviness/lump in hypochondrium.
d. Tender hepatomegaly.
• Anchovy sauce pus in the centre of abscess.
• Diagnosis :
a. USG
b. WBC, Liver enzymes.
Treatment : Metronidazole (Including asymptomatic patients to prevent

transmission).
Erythrophagocytosis
Trophozoites of E. histolytica Flask-shaped ulcers Quadrinucleate cyst of

E.histolytica.
Free living amoeba :
1. Naegleria fowleri/Brain-eating amoeba :
Human : Accidental, dead-end host.
Morphological forms : Cyst, amoeboid & flagellate trophozoite (Infective form).
MOI : Nasal mucosa (swimming) Cribriform plate Brain.
Disease : Primary amoebic meningoencephalitis.
a. Acute & highly fatal condition.
b. CSF examination similar to bacterial
meningitis.
Diagnosis :
a. PCR.
Lobopodia of amoeboid trophozoite
b. Antigens in CSF. of Naeglaria fowleri on culture.
c. Microscopy : Amoeboid trophozoite.
d. Culture of CSF : Non nutrient agar + Lawn culture of E.coli.
Treatment : IV & intrathecal Amphotericin B (Cornerstone).

94 08 Microbiology

2. Acanthamoeba :
Morphological forms : Cyst & trophozoite (Both are infective forms).
Human : Accidental & dead-end host.
MOI & disease :
a. Via corneal trauma keratitis.
b. Skin/respiratory tract Granulomatous
amoebic encephalitis (Immunodeficiency is a risk
factor & slow onset).
Trophozoite of
Diagnosis : Acanthamoeba in culture
a. Brain biopsy/PCR/antigen test. (Spine-like acanthopodia).
b. Microscopy/Culture of tissue (Non nutrient agar + lawn culture of
E.coli).
3. Balamuthia mandrillaris :
Morphological forms : Cyst & trophozoite.
MOI & disease : Skin/respiratory tract
Granulomatous amoebic encephalitis.
(Immunodeficiency is a risk factor & slow onset).
Trophozoite of Balamuthia in culture

(Long pseudopodia).
Flagellates 00:11:48
1. Giardia lamblia/Grand old man of intestine :

M/C parasitic infection & M/C parasite in stool sample.
Morphological forms : Cyst (Infective form) & trophozoite (disease form).
Host : One (Human).
Habitat : Upper small intestine (Duodenum & upper jejunum).
M.O.I : Ingestion of quadrinucleate cyst (Small ID50).
Disease : Trophozoites attach to mucosa & cause :
a. Asymptomatic colonization in 50%.
b. Acute self-limited diarrhoea with malaise, cramping & bloating.
c. Chronic diarrhoea malabsorption & steatorrhoea (Fatty stools).
Diagnosis : Stool sample.
a. PCR (Gold standard)/Antigen.
b. Microscopy (for trophozoite & cysts) : Falling leaf motility of trophozoite.
c. Culture of stool (Diamond’s medium).
d. Enterotest/string test (not in use).
Treatment : ----- Active space -----

a. Tinidazole once Or
b. Metronidazole x 7 days.
H & E stain : Duodenal biopsy in giardiasis

(Sickle shaped trophozoites).
Diarrhoea causing agents with small ID50 Invasive Diarrhoea

E. histolytica V BE SECSEY group
G. lamblia Vibrio parahaemolyticus
Cryptosporidium Balantidium coli
Entamoeba histolytica
Shigella
Bacteria Shigella
EHEC
EIEC
Campylobacter
Salmonella
EHEC
Yersinia enterocolitica
2. Trichomonas vaginalis :
Morphological forms : Trophozoite only (Infective form).
Host : One (Human : Obligate parasite).
M.O.I : Sexual transmission (M/C parasitic STD).
Disease :
• Asymptomatic colonization in 10-50%.
• Urethritis in males.
• Vaginitis :
a. Frothy, foul greenish discharge, vaginal pH>4.5.
b. Dyspareunia , dysuria.
c. O/E : Strawberry cervix (D/t microulcerations).
• Dx : Microscopy, Culture, PCR.
Treatment : Metronidazole/tinidazole once.
3. Trypanosoma cruzi/Chagas disease : Trophozoites of T. vaginalis

M/C cause of infectious myocarditis. (Jerky/wobbling/twitching/
Reported : Central & South America. rotatory motility).
Definitive host : Humans, opossum, armadillo.
Morphological form : Metacyclic trypomastigote (Infective form).
Vector : Reduviid/Triatoma bug (Kissing bug).
96 08 Microbiology

M.O.I : Feces of bug, across placenta, blood, organ transplantation.
Disease :
• Asymptomatic : Most patients.
• Acute Chagas disease :
a. Chagoma : Painless swelling at the site of entry of trypomastigote.
b. Romana sign : Periorbital chagoma.
c. High grade fever, lymphadenitis, carditis, meningitis, encephalitis.
d. Diagnosis : Blood sample & lymph node aspirate.
i. PCR, antigen.
ii. Microscopy of blood sample : Circulating trypomastigote.
Lymph node sample : Resting amastigote.
iii. Culture of buffy coat & LN biopsy.
• Chronic disease (10-30%) :
a. Dilated cardiomyopathy, megaesophagus, megacolon.
b. Diagnosis : Serology (ELISA).
c. Xenodiagnoses : Reduviid bug.
Treatment :
a. Benznidazole (Or)
b. Nifurtimox.
Triatomine bug T. cruzi trypomastigote in a Romana sign

thin blood smear stained with
giemsa stain.
4. Trypanosoma brucei/African sleeping sickness (SS) :
West African SS (WASS) East African SS (EASS)
Definitive host Humans only Humans, domestic & wild mammals
Organisms T.b.Gambiense T.b. Rhodesiense
(WAGS EARS) (Rapidly progressing).
Vector : Glossina/tsetse fly.
M.O.I : Bite, across placenta, blood, organ transplantation.

Disease : ----- Active space -----

a. Bite site : Painful trypanosomal chancre or swelling.
b. Hemolymphatic stage : Fever, LNAP, rash, edema, carditis.
Winterbottom sign : Posterior cervical LNAP (Often in WASS).
c. Neurological stage :
Alteration of sleep rhythm, mental confusion, mania, depression, coma.
Diagnosis : Blood/CSF microscopy (For trypomastigote), culture (NNN media).
Note : Amastigote not seen in microscopy.
Treatment :
WASS EASS
Hemolymphatic stage Pentamidine Suramine
Neurological stage Eflornithine Melarsoprol
T.brucei trypomastigote in Tsetse fly

a thin blood smear
5. Leishmania :
Morphological form :
a. Promastigote (Infective form) & amastigote (Seen in man).
b. Kinetoplast present in both the forms.
Visceral leishmaniasis/kala azar :
Agent : L. donovani complex.
Endemic areas : Bihar, Orissa, Rajasthan.
Definitive host : Humans.
Vector : Phlebotomus argentipes (Sand fly).
M.O.I : Bite, transplacental, blood, organ transplantation.
Pentad of kala azar :
1. Prolonged fever
2. Progressive weight loss
3. Pronounced hepatosplenomegaly (HSM)
4. Pancytopenia
5. Hypergammaglobulinemia

98 08 Microbiology

Diagnosis :
• Sample : Spleen (Most sensitive) > bone marrow biopsy > lymph node > liver
biopsy > blood buffy coat.
• Tests :
a. PCR, antigen, culture (NNN media).
b. Microscopy (LD bodies/amastigote in macrophages) ; Localizes to
Kupffer cells.
c. Serology :
Specific : Rk39 antigen.
Non specific : Chopra’s antimony test/Napier’s aldehyde test.
Treatment :
a. DOC outside east Africa : Liposomal amphotericin B.
b. Post kalaazar dermal leishmaniasis (PKDL) : Miltefosine orally.
Cutaneous leishmaniasis :
Old world : L tropica, L. major.
New World : L. mexicana, L. amazonenesis.
Definitive host : Humans, dogs.
Vector : Phlebotomus sergenti (Sand fly).
M.O.I : Bite .
Disease : Cutaneous leishmaniasis/Baghdad boil/Aleppo sore/chiclero ulcer.
Diagnosis : PCR, antigen, culture, microscopy (LD body).
Treatment : Sodium stibogluconate (Pentavalent antimonial compound).
LD body/amastigote Promastigotes in NNN Sand fly

in blood smear stained medium
with Giemsa.
Balantidium coli 00:32:47
Largest protozoa of man.

Morphological forms : Ciliated trophozoite (Invasive form), binucleate cyst
(Infective form).
Host : One (Human).

Animal reservoir : Pigs. ----- Active space -----

Habitat : Large intestine. Cilia
M.O.I : Ingestion of binucleate cyst.
Disease :
Macronucleus
a. Asymptomatic in 50-80%.
b. Ciliate dysentery (Invasive diarrhea).
Diagnosis : Stool (Microscopy, PCR). Trophozoite of Balantidium coli
Treatment :
a. TOC : Tetracycline.
b. Metronidazole/iodoquinol.
Coccidean protozoa 00:34:47
Cryptosporidium hominis & C. parvum, Cystoisospora bellii, Cyclospora

cayatenensis :
Self limited diarrhea in healthy people.
Intractable & unrelenting diarrhea in HIV/immunodeficient.
Host : One.
a. Humans (C. hominis, C. bellii, C. cayatenensis).
b. Mammals (C. parvum).
Habitat : Small intestine .

Mode of infection : Ingestion of sporulated oocyst (Infective form).
Disease :
a. Watery diarrhoea.
b. Autoinfection : Cryptosporidium.
Diagnosis :
• Sample : Stool.
• Tests :
a. Modified Z.N/kinyoun staining : Partially
acid-fast oocysts.
b. UV light microscopy for fluorescence. Acid fast oocyst on stain.
c. PCR for gene amplification.
Important points :
• M/C cause of diarrhoea in HIV : Cryptosporidium.
• Small ID50 : Cryptosporidium.
• Internal and external autoinfection : Cryptosporidium.
• Zoonotic infection : C. parvum.
100 08 Microbiology

• Unique extra cytoplasmic location (Parasitophorous vacuole in the microvillus)
: Cryptosporidium.
Cryptosporidium Cyclospora Cystoisospora
Size of oocysts 4-5 microns 8-12 microns 20-33 microns
Sporozoites in oocysts in
Yes No
feces
Autofluorescence in oocysts - ++ +
Treatment Nitazoxanide Cotrimoxazole
Nematozoans 00:40:56
1. Plasmodium :
Host : Two.
a. Definitive host : Female anopheles.
b. Intermediate host : Humans.
Habitat in human : Hepatocytes & RBCs.
Mode of infection : Sporozoites via bite, blood transfusion, across placenta
Blood forms : Trophozoites.
Pathogenesis :
Life cycle of Plasmodium

Secondary erythrocytic schizogony : Seen in P. vivax & P. ovale d/t activation of
hypnozoites Relapse of malaria.

• Microscopy (Gold standard) :
a. Thick smear : Presence of & quantification of parasitemia.
b. Thin smear : For species identification.
Note : Romanowsky stains (Giemsa stain/Wright stain/Leishman stain) &
Jaswant Singh & Bhattacharya (JSB) stain are used.
• Rapid tests : Immunochromatographic tests.
a. Aldolase/LDH : Pan malarial antigen.
b. HRP-II (Histidine rich protein) : Plasmodium falciparum.
• QBC (Quantitative buffy coat)/Microhematocrit method.
• PCR (Most sensitive). Rapid test kit
• Culture on RPMI medium.
Recrudescence : Low level of parasitemia (d/t inadequate treatment)
Multiplication of parasite in blood stage Infection.
Complications of Malaria :
a. Cerebral malaria.
b. Nephropathy : Seen in P. malariae d/t immune complex deposition.
Note :
a. Duffy antigen (Receptor) negativity Resistance to P. vivax.
b. Protection against severe malaria : Sickle cell trait, HbF, thalassemia,
G6PD deficiency.
c. P. knowlesi is a zoonotic malaria (Reservoir : Chimpanzee).
P. falciparum P. vivax P. ovale P. malariae P. knowlesi
Malaria Malignant Benign Ovale Quartan Quotidian
tertian tertian malaria malaria malaria
malaria malaria
Erythrocytic 48 hours 48 hours 48 hours 72 hours 24 hours
schizogony
RBC Younger cells Retics & Retics Older cells Younger
preference (all ages) cells up to cells (all
2 weeks old ages)
Vacuoles Maurer’s Schuffner’s Schuffner’s Ziemann’s
Hemozoin Black Yellow Dark brown Brown Dark
brown black brown
Incubation 12 days 13-17 days 13-17 days 28-30 10-12 days
period days

102 08 Microbiology

Peripheral blood smear :
P. falciparum P. vivax & ovale P. malariae
Crescent shaped gametocyte Ameboid gametocytes Band-shaped trophozoite
Accole form Enlarged RBCs (All stages seen)
2. Babesia :
Host : Two .
a. Definitive host : Hard tick (Vector).
b. Intermediate host : Mammals.
c. Accidental & dead end host : Man.
Habitat in human : RBCs.
Note : No hepatocyte stage is seen in Babesia.
M.O.I : Sporozoites (Infective form) via bite, blood transfusion, across placenta.
Blood forms : Trophozoites.
Disease : Sporozoites invade RBCs.
a. Fever, chills, sweats, Hemarthrosis, myalgias.
b. Severe babesiosis : Seen in asplenia, CD4,
B. divergens, transplant recipient patients.
Diagnosis :
a. PCR, antigen.
b. Microscopy : Maltese cross (Tetrad form).
Treatment : Maltese cross
a. Moderate disease : Atovaquone PO + azithromycin PO X 7-10 Days.
b. Severe disease : Clindamycin PO + quinine PO X 7-10 Days.
c. RBC transfusion for B. divergens.

3. Toxoplasma gondii : ----- Active space -----
Hosts : Two.
a. Definitive host : Cat.
b. Intermediate host (I.M.H) : Birds, rodents, sheep.
c. Accidental host : Man.
Mode of infection :
1. Poorly cooked meat of I.M.H : Tissue cyst (Bradyzoites).
2. Cat feces : Oocysts.
3. Blood transfusion/organ transplant/transplacental :
Endodyogeny :
Tachyzoites. Unique replication method
of tachyzoites of
toxoplasma.
Toxoplasma gondii
1° Healthy Immunodeficient patient

Reactivation of cysts/1° Persistence of tachyzoites
Infections : Encephalitis (M/C) : Chorio-retinitis

a. Asymptomatic • CT & MRI : multiple ring Necrotizing lesions :
b. Painless cervical LNAP enhancing lesions multifocal/bilateral
c. Generalized LNAP • M/C cause of intracere-
Spontaneous resolution in b & c bral mass lesions in HIV Pneumonitis

104 08 Microbiology

Diagnosis :
• Sample : CSF/BAL/Aqueous humor.
• Tests :
a. PCR, Antigen tests.
b. Giemsa stain to demonstrate tachyzoites.
c. Cultured only cell lines.
d. Serology : IgM & IgG using Sabin Feldman dye test (Gold standard).
Sabin Feldman dye test : Complement based test.
Live tachyzoites + Serum + Complement = Lysis of RBC (If IgG+ present).
+
Methylene blue = No uptake of dye
Inference : IgG Ab present.
Treatment : Pyrimethamine + folinic acid + sulfadiazine.
Toxoplasma gondii in pregnancy :
• Serology :
a. IgM & IgG : Both +ve in recent infection.
b. Igm -ve + IgG +ve/High avidity IgG : Remote infection.
• Treatment :
a. Acute infection : Spiramycin till 18 weeks.
b. Pyrimethamine + Folinic acid + Sulfadiazine till delivery.
Transmission with duration.
Severity with duration.
• Congenital toxoplasmosis : 3 C’s. 1. Chorioretinitis.
2. Hydrocephalus.
3. Intracranial calcification.

Helminths 00:02:28
Cestodes Trematodes Nematodes

Flat, tape like. Flat, leaf like. Roundworms.
Segmented. Unsegmented. Unsegmented.
Hermaphrodites (Monoecious). Diecious.
Incomplete alimentary Complete alimentary
Absent alimentary canal.
canal. canal.
Trematodes 00:03:56
Common features except for Schistosoma :

• Definitive host : Mammals.
• Intermediate host :
1. 1st intermediate host : Snails.
2. 2nd intermediate host : Fresh water fish/
plants.
• Mode of infection : Ingestion of metacercariae in
poorly cooked fish/unwashed water plants.
Adult fluke
• Infective form : Metacercariae.
• Operculated egg.
Classification of trematodes based on habitat :

1. Liver flukes.
2. GIT flukes.
3. Lung flukes (Paragonimus westermanii/ Operculated egg which open
Oriental lung fluke). to release miracidium in
4. Blood flukes (Schistosoma species). water.
Toc for trematodes : Praziquantel except Fasciola spp. DOC is Triclabendazole.

106 09 Microbiology
----- Active space ----- Liver flukes :

Fluke Definitive 1st inter- 2nd intermediate Pathogenicity Diagnosis
host mediate host
host
Fasciola • Asymptomatic
hepatica. Sheep, (M/c).
Fresh water
cattle, • Cholangitis.
Fasciola plants.
man. Stool for
gigantica. • Cholecystitis.
Snail. operculated
• Cholelithiasis.
Clonorchis eggs.
Cat, dog, • Biliary cirrhosis.
sinensis. Fresh water fish.
man.
Opisthorchis.
Cholangiocarcinoma : Seen with Clonorchis sinensis & Opisthorchis viverrini.
GIT flukes (Intestinal distomiasis) :

1st 2nd
Definitive
Fluke intermediate intermediate Pathogenicity Diagnosis
host
host host
Fasciolopsis buski • Asymptomatic
Man, pig. (M/c).
(Small intestine).
• Abdominal Stool for
Fresh water
Snail. pain. operculated
Gastrodiscoides plants.
• Mucous eggs.
hominis Man, birds.
diarrhoea.
(Large intestine).
Lung flukes/endemic hemoptysis/pulmonary distomiasis :

1st inter- 2nd inter-
Definitive
Fluke mediate mediate Pathogenicity Diagnosis
host
host host
• Asymptomatic
• CXR findings :
Crusta- (M/c). Nodular opacities
Paragonimus Man, pig, ceans : • Cough with & pleural effusion.
Snail.
westermanii. cat, rat. Crab/cray- rusty brown • Sputum & stool
fish. sputum. for operculated
• Frank/chronic eggs.
haemoptysis.
Paragonimiasis Vs TB (Clinically) :
1. Lack of fever.
2. Eosinophilia.
Blood flukes (Bilharziasis) : ----- Active space -----

Schistosomes :
They have separate sexes.
Only one intermediate host : Snail.
Mode of infection : Penetration of skin.
Infective form : Cercariae.
Non operculated eggs with spines.
Inter-
Definitive
Fluke Habitat mediate Pathogenicity Diagnosis
host
host
Schistosoma Inferior • Cercarial dermatitis.
Man, pig. • Katayama fever.
mansoni. mesenteric
plexus.
In chronic disease : Stool for non
• Hepatosplenic
Superior schistosomiasis. operculated.
Schistosoma
mesenteric • Egg embolism
japonicum.
plexus. (Maximum :
Snail. Japonicum).
Man, • Cercarial dermatitis.
birds. • Katayama fever
Schistosoma (Rare). Urine
Vesical
haematobium. • Hematuria. for non
plexus.
• Hydroureter, operculated..
hydronephrosis.
• SCC of Bladder.
Egg of S. haematobium Egg of S. mansoni in Egg of S. japonicum in

in urine stool stool
Terminal spine. Lateral spine. Lateral knob.
Life cycle of Schistosomes.

108 09 Microbiology

Note : Schistosomes are not indigenous to India, always h/o of travel to endemic
areas will be present.
Katayama fever : Occurs first time when females lays eggs due to immune
response against eggs → C/f : Fever with chills & hepatosplenomegaly.
Nematodes 00:20:30
Infective form : L3 larva.

Intestinal nematodes Tissue nematodes
One host. Two hosts.
Mode of infection : Ingestion/skin
Mode of infection : Arthropod bite.
penetration.
Oviparous/ovoviviparous. Viviparous.
• Strongyloides. • Wuchereria.
• Ascaris. • Brugia.
Small intestine.
• Necator. • Onchocerca.
• Ancylostoma. • Loa.
• Trichinella.
• Toxocara.
Large intestine.
• Enterobius.
• Trichuris.
Small intestinal nematodes :

1. Strongyloides stercoralis :
Mode of
Special points Pathogenicity Diagnosis
infection
• Opportunistic Larval :
infection. • M/c : Asymptomatic.
• Parthenogenic. • Larva currens. Stool for
• Ovoviviparous. • Pneumonitis. rhabditiform
• Internal & external Skin penetration • Hyperinfection. larva
autoinfection. by filariform • Disseminated infection. (Baerman
larva(L3). funnel, harada
• No eggs in stools. Adult worm :
• Free living in soil. mori, agar plate
• Abdominal pain. culture).
• Diarrhoea/constipation.
• Malabsorption.
Rx :
• DOC Ivermectin x 2 days.
• If disseminated : Ivermectin x 14 days.

Rhabditiform larva (L1)

in stool.
Fig : Life cycle of Strongyloides.
2. Ascaris/roundworm :
Mode of
infection
• M/c helminth infection • Asymptomatic (M/c).
in the world. • Loffler’s syndrome :
• Longest nematode of • Malnutrition.
Stool for
human intestine. Ingestion of eggs • Vit A deficiency.
bile stained
with larva (L3). • Intussusception.
eggs.
• Intestinal obstruction.
• Appendicitis.
• Pancreatitis.
Rx : Albendazole 400 mg once/ Mebendazole x 3 days/ Pyrantel palmoate x 3 days.
Fertilized egg of Ascaris

Bile stained
Single unsegmented ovum.
Unfertilized egg of Ascaris

Bile stained
Life cycle of Ascaris. Atrophied ovum.
110 09 Microbiology

Visceral larva migrans :
• Due to accidental ingestion of eggs of cat/dog roundworms (Embryonated
egg with L3 larva).
• Caused by :
1. Toxocara canis (Dogs).
2. Toxocara cati (Cats).
3. Ascaris suum.
4. Baylisascaris procyinis.
• C/f :
a. M/c asymptomatic.
b. Eosinophilic granulomas.
c. Hepatosplenomegaly.
d. Neuroretinitis.
e. Eosinophilic meningitis .
3. Hookworms :
• Necator americanus (New world hookworm).
• Ancylostoma duodenale (Old world hookworm).
Mode of infec-
tion
M/c : Asymptomatic.
• Ground itch.
Skin penetration • Pneumonitis. Stool for non
Special mouth parts :
by filariform larva • Abdominal pain. bile stained
Teeth/cutting plates.
(L3). • Dyspepsia. eggs.
• Microcytic hypochromic
anemia.
Rx : Albendazole 400mg once/mebendazole � 3 days/pyrantel pamoate � 3 days.
Note : M/c cause of loffler’s syndrome : Ascaris.
Eggs of hookworm
Non bile stained egg
Segmented ovum
4 - 8 blastomeres.
Life cycle of hookworms.
Cutaneous larva migrans/creeping eruption : ----- Active space -----

• Caused by non human hookworms.
1. Ancylosoma brasiliensis (M/c).
2. Ancylosoma caninum.
3. Uncinaria stenocephala.
4. Bunostomium phlebotomum.
5. Gnathostoma spinigerum.
• C/f : Vesicular erythematous eruption, slowly Cutaneous larva migrans.
progressive (Few cm/day).
Note : In larva currens, progression of rash is rapid (5 - 10 cm/hr).
Large intestinal nematodes :

1. Enterobius vermicularis : Pin worm/Seat worm.
Mode of infection Pathogenicity Diagnosis
• Fomites. • Asymptomatic (M/c).
• Autoinfection . • Perianal & perineal
Perianal swab (NIH/scotch
• Retro infection . pruritis.
tape) for non bile stained
• Oro genital sex. • Enuresis.
eggs.
• Abdominal pain.
• Vaginitis & urethritis.
Rx : Albendazole/Mebendazole/Ivermectin once → Repeat after 2 wks.
Egg of Enterobius
vermicularis
Planoconvex shaped.
Fig : Life cycle of Enterobius vermicularis.

112 09 Microbiology
----- Active space ----- 2. Trichuris trichiura : Whip worm.

Mode of infec-
Pathogenicity Diagnosis
tion
• Asymptomatic (M/c).
• Colitis :
1. Tenesmus.
Ingestion of 2. Abdominal pain. Stools for bile
embryonated egg. 3. Mucoid/bloody diarrhoea. stained eggs.
• Rectal prolapse (Coconut cake rectum : Due to
intense inflammation).
• Iron deficiency anemia.
Rx : Albendazole x 3 days, mebendazole x 3 days.
Egg of Trichuris trichiura.

Barrel shaped with bipolar mucus plugs.
Tissue nematodes :
Wuchereria & Brugia :
Mode of
Nematode Vector Pathogenicity Diagnosis
infection
M/c : Asymptomatic • Microfilaria (L1
Adult worms : larva) in blood
• Acute filarial fever : Self btw 10 pm - 2
limiting fever with chills. am (Nocturnal
Wuchereria • Lymphatic obstruction. periodicity).
bancrofti. Culex. • Quantitative
(M/c) Hypersensitivity (HSR) to buffy coat/micro
microfilaria : haematocrit tube
Bite of
• Tropical pulmonary eo- method.
mosquito
sinophilia (TPE) : Noc- • Ag detection.
(L3 larva).
turnal attacks of cough, • PCR.
wheezing & dyspnea.
• Occult filariasis : hepat- TPE :
Brugia malayi
osplenomegaly. • Eosinophilia
(Eastern India,
• IgE .
far east & Mansonia.
• Microfilarial Ab’s .
South East
• No microfilaria in
Asia).
blood.
Rx : DEC x 12 days.

Wuchereria. Brugia.
Life cycle of Wuchereria/Brugia.

114 09 Microbiology

Cestodes 00:45:11
Common features :
• Definitive host : Humans (Small intestine) except Echinococcus (Dog/wolf).
• Intermediate host : One/two/none.
• Mode of infection : Ingestion.
• Infective form : Variable.
• Appearance :
Grooves Diphyllobothrium latum.
Scolex No rostellum Taenia saginata.
Suckers Rostellum with hooklets
Rostellum with no hooklets Hymenolepsis diminuta.
Hooklets
Scolex/head
Proglottids
/segments
Suckers with Suckers Grooves

rostellum
Diphyllobothrium latum :
Fish/broad tape worm.
Longest tape worm infecting man.
1st inter-
Definitive
mediate 2nd intermediate host Pathogenicity Diagnosis
host
host
• Asymptomatic
(M/c).
Man & fish • Abdominal pain. Stool for
Fresh water fish (With
eating Cyclops. • Diarrhoea. operculated
plerocercoid larva).
mammals. • Obstruction. eggs.
• Vit B12 deficiency
anemia.
Egg of D. latum
Operculated.
Knob at abopercular end.

Hymenolepis : ----- Active space -----
Definitive
Cestode Intermediate host Pathogenicity Diagnosis
host
Hymenolepis nana In direct cycle : None. Stool for

(Dwarf tape worm) M/c : non bile
Smallest worm In indirect cycle : Moths, Asymptomatic. stained
infecting man. Man & beetles & fleas. • Anorexia. eggs.
Hymenolepsis rodents. • Abdominal Stool
diminuta pain. for bile
Moths, beetles & fleas. • Diarrhoea.
(Rat tape worm) stained
Rare infection. eggs.
Rx : Niclosamide x 1 wk/praziquantel once.
External Internal
autoinfection autoinfection
Life cycle of H. nana
Egg of H. nana Egg of H. diminuta

Non bile stained Bile stained
Hexacanth oncosphere, Yolk granules & Hexacanth oncosphere, No yolk
4-8 polar filaments. granules & polar filaments.

116 09 Microbiology

Taenia species :
Intermediate
Cestode Definitive host Pathogenicity Diagnosis
host
Taenia saginata Man
(Beef tape worm) (Ingestion of Cows. • Asymptomatic
4-6 mts. cysticercus bovis). (M/c).
Stool for
Taenia solium • Anorexia.
Man eggs.
(Pork tape worm) Pigs (Poorly • Abdominal pain.
Ingestion of cystic- • Diarrhoea.
2-3 mts. cooked).
ercus cellulosae).
Egg of taenia spp.

Hexacanth oncosphere
with radially striated
embryophore.
Cysticercosis :
Caused due to autoinfection of taenia solium.
Life cycle of taenia solium.
Neurocysticercosis :
M/c parasitic infection in the world.
M/c site : Intracerebral hemisphere.
C/f :
• M/c : New onset partial seizure.
• Hydrocephalus.
• Meningitis.
Dx : Imaging + serology (ELISA, western blot). Neurocysticercosis.

Echinococcus species : ----- Active space -----
Definitive Interme-
Cestode Pathogenicity Diagnosis
host diate host
Echinococcus Dogs Hydatid disease
Sheep,
granulosus (Eggs • M/c : Asymptomatic.
goats,
(Dog tape worm). in dog • M/c site : Liver.
Humans. Imaging +
faeces). • 2nd m/c : Lung.
serology
• M/c : Asymptomatic. (ELISA,
• Echinococcosis/alveolar hy- western
Echinococcus Wolves, datid disease. blot).
Rodents.
multilocularis. foxes, dogs. • Multi cystic hydatid cyst :
Thin peri cyst → spread
rapidly.
Hydatid cyst
Life cycle of Echinococcus granulosus.

118 09 Microbiology

Man is intermediate host in : ToP BET.
a. Toxoplasma.
b. Plasmodium.
c. Babesia.
d. Echinococcus.
e. Taenia solium.
Non bile stained eggs :

A HEN laid non bile stained eggs.
a. Ancylostoma duodenale.
b. H. nana.
c. E. vermicularis.
d. N. americanus.
Parasites acquired by skin penetration :

SCANS
a. Schistosomes.
b. Cutaneous larva migrans.
c. A. duodenale.
d. N. americanus.
e. Strongyloides.

Fungi : General characteristics 00:01:14
Fungi are eukaryotes.

Structure :
Morphology :
Cryptococcus spp.,
Yeast
Saccharomyces cerevisiae (Bakers/brewers yeast)
Yeast like :
Candida spp., Malassezia spp
Pseudohyphae
Mold :
Rhizopus, Mucor, Conidiobolus, Basidiobolus
Aseptate
Hyaline (melanin -) : Aspergillus, Fusarium, Acremonium,
Penicillium.
Mold : Septate
Phaeoid (melanin +) : Madurella, Exophiala, Cladosporium,
Phialophora, Fonsecaea, Cladophialophora.
Sporotrichosis, Talaromyces, Coccidioidomycosis,
Dimorphic
Blastomycosis, Paracoccidioidomycosis, Histoplasmosis
Dimorphic :
At 37 °C : Yeast. Mnemonic :
At room temperature : Hyaline septate mold. BodY → Yeast.
Never pigmented. RooM → Mold (Hyaline septate).

120 10 Microbiology

Microscopy :
Stains Fungi stained Special points
Gomori/Groco
methenamine silver All fungi stained black Best fungal stain
stain
Periodic acid Schiff
All fungi stained magenta -
(PAS)
Masson fontana silver
Pigmented/phaeoid -
stain
Calcofluor white - Fluoresecent stain
Splendore hoeplii
Hematoxylin & eosin Sporotrichosis phenomenon/asteroid
body
Giemsa Histoplasma -
Capsule of Cryptococcus
Mayer mucicarmine
neoformans/gattii in -v
stain
tissue samples
GMS stain showing Cryptococcus PAS stain showing Histoplasma
Wet mounts :
Negative stain (India ink/Nigrosin) Cryptococcus in CSF/blood specimens
Lacto phenol cotton blue stain (LPCB) Culture morphology
KOH mount Fungi in skin/hair/nail

Culture : ----- Active space -----

pH : 5.4-5.6
Sabouraud dextrose agar
Gentamicin/chloramphenicol to inhibit bacterial
(M/C)
growth
Brain heart infusion agar
Dimorphic fungi
(BHIA) + 5% blood
Niger seed/bird seed
Pathogenic cryptococcus produce brown colonies
agar
CHROM agar Identify species of candida based on colour
Corn meal/rice meal agar Nutritionally deficient medium
Chlamydospores of C. albicans & C. dubliniensis
Tests :
All invasive mycoses (Except cryptococcosis &
β 1,3 D glucan (Serum)
mucormycosis)
Galactomannan (Serum) Invasive aspergillosis
To identify Trichophyton mentagrophytes/
Hair perforation
Microsporum canis
Urease Cryptococcus species (Pink color)
Canavanine glycine
Cryptococcus neoformans (Negative) vs
bromothymol blue (CGB)
gattii (Positive)
test
Superficial mycoses 00:15:52
Pityriasis/Tinea versicolor :
Malassezia furfur & M. globosa.
Yeast-like fungus.
Lipophilic.
Seen on chest, neck & trunk.
Yellow-green fluorescence under Wood’s
lamp.
Rx : Topical ketoconazole/terbinafine.
Hypo/hyperpigmented/erythematous
non itchy macules surrounded
by soft/furfuraceous scales.

122 10 Microbiology
Spaghetti meatball appearance/

SDA with olive oil
banana grapes appearance
Tinea/ring worm/dermatophytosis :
Tissue specimens : Hyaline septate molds + arthrospores.

SDA agar : Hyaline septate molds + macroconidia & microconidia.
Dermatophytes Macroconidia Microconidia Affects
Trichophyton Pencil ++ Skin, hair, nail
Epidermophyton Club - Skin, nail
Microsporum Boat/spindle/fusiform + Skin, hair
Human to human
Anthropophilic T. rubrum, T. schonleinii, E. floccosum
transmission
Zoophilic Reservoir : Animals T. verrucosum, T. mentagrophytes
Geophilic Reservoir : Soil T. terrestre
LPCB mount :
Trichophyton spp. Epidermophyton spp. Microsporum spp.
T.rubrum
Tinea cruris Tinea pedis Tinea unguium

Jock itch/Dhobi itch Athlete’s foot
Inflammatory types of T. capitis
Hair perforation test Kerion (boggy swelling) Favus (yellow crusts)

+ve for T.mentagrophytes T.verrucosum T.schoenlenii >
& M.canis M.canis T.violaceum
T. mentagrophytes
Subcutaneous mycoses 00:23:58
Chromoblastomycoses :
Dematiaceous (Phaeoid) fungi :
• Fonsecae spp.
• Cladophialophora.
• Phialophora.
• Rhinocladiella.
• Chromomycosis. Copper penny app.
Causes : Chromomycosis/Verrucous dermatitis.
Trauma → Chronic granulomatous disease of skin & subcutaneous tissues only.
Lesions : Nodular, tumoral (cauliflower like), verrucous, plaques, cicatricial or
mixed.
Skin scrapings (KOH) : Medlar/muriform/sclerotic/copper penny bodies.
Rx :
Small lesions : Surgical excision/cryotherapy/photocoagulation.
Large lesions : Surgical removal + Itraconazole/Terbinafine X 3-24 months.

124 10 Microbiology
Mycetoma/Maduramycosis :
Acquired through trauma, chronic granulomatous disease of skin & underlying

tissues (Including muscles & bones).
C/F : Swelling + sinuses + granules (Collection of organisms).
Type of
Organisms Granules Rx
organism
Small lesions :
Aerobic acti- Off-white/ Trimethoprim
Nocardia,
Actinomy- nomycetes/ yellow (TMP) alone.
Actinomadura,
cetoma branching granules Large lesions :
Streptomyces
bacteria TMP +
Amikacin.
Phaeoid
(Madurella/ Brown
Exophiala) Surgical
Eumyce-
Fungi Hyaline debulking with
toma
(Pseudallescheria, azoles
Off-white
Acremonium,
Aspergillus)
M/C cause of mycetoma in India : Actinomadura madurae.
M/C cause of eumycetoma in India : Madurella mycetomatis.
Mycetoma/madura foot
Opportunistic fungal infections 00:30:14
Candida :
Yeast like fungus.

Forms yeast & pseudohyphae.

Types : ----- Active space -----

1. C. albicans.
2. Non albicans spp. :
• C. dubliniensis.
• C. glabrata
• C. auris Often resistant to azoles
• C. krusei
Tests :
1. Germ tube test/Reynolds-Braude phenomenon +ve +ve for C. albicans &
2. Corn/rice meal agar : Chlamydospores C. dubliniensis.
3. CHROM agar :
• Light green : C. albicans.
• Dark green : C. dubliniensis.
Note : C. auris (Candida super bug) is resistant to all
types of antifungals.
Germ tube test.
Candidiasis :
Mucocutaneous Invasive
Neutropenia, steroids,
Risk factors HIV, DM, pregnancy hematological ca, prolonged
antibiotics
Intertrigo, thrush, Vulvo
Vaginal candidiasis, Candidemia, meningitis,
Diseases
oesophageal candidiasis, pneumonia, hepatitis
diaper rash, onychomycosis
Topical /oral Amphotericin B, azoles &
Treatment
azoles echinocandins
CHROM agar showing Chlamydospores

different Candida spp

126 10 Microbiology

Aspergillus :
Hyaline septate mold.

Acute angled/dichotomous branching (branching at 45 degrees), includes :
• A. fumigatus.
• A. flavus.
• A. niger.
• A. terreus (resistant
to all azoles).
It causes :
• Allergic Broncho Pulmonary Aspergillosis
(ABPA).
• Sinusitis.
• Otomycosis (A. niger).
• Keratitis (Trauma with vegetative A. fumigatus
matter).
• Fungus ball (A. fumigatus).
• Aflatoxins (A. flavus & A. parasiticus).
Invasive pulmonary aspergillosis :

Risk factors : Neutropenia, HIV pts, transplant A. flavus
recipients.
M/C organism : A. fumigatus.
Diagnosis : Serum galactomannan; β 1,3 D
glucan.
Biopsy : Dichotomous branching.
Treatment :
• DOC : Voriconazole.
• Other drugs : Isavuconazole, liposomal Aspergillus (acute angled branching)
amphotericin B.

Mucorales : ----- Active space -----
Hyaline aseptate mold.

Wide angled branching (90 degrees).
Bread molds.
They include :
• Rhizopus (M/C).
• Mucor.
Asexual spores : Sporangiospores.
Sexual spores : Zygospores.
It causes mucormycosis/zygomycosis/phycomycosis.
Angioinvasion +.
Risk factors : Uncontrolled DM, steroids, iron over-
load, deferoxamine therapy, transplant recipients
(TR).
M/C type : Rhino cerebral/Rhino ocular/Rhino oculo Aseptate, ribbon like hyphae
cerebral mucormycosis.
Note : Cutaneous mucormycosis occurs in immunocompetent people.
Diagnosis : Biopsy, Serum β 1,3 D glucan negative.
Rx : Wide surgical resection + Amphotericin B.
Pneumocystis jirovecii :
Ergosterol absent→ Antifungals are not effective.

Trophozoites & cysts forms (Like protozoa).
Non cultivable on cell free media.
Risk factors : HIV (CD4 <200), transplant recipients, immuno deficiency states.
Bilateral interstitial pneumonia (plasma cell pneumonia/PCP) : Infilitration starts
at perihilar region & spreads peripherally.
Diagnosis :
BAL or induced sputum.
Serum β 1,3 D glucan +ve.
Rx :
DOC : Cotrimoxazole.
Cysts stained by GMS
(hat/ping pong ball appearance).

128 10 Microbiology

Cryptococcus :
Only pathogenic fungus which has a capsule.

C. neoformans C. gatti
Capsule serotypes A&D B&C
Spreads through soil
Pigeon droppings Eucalyptus
contaminated by
Urease +ve +ve
CGB test -ve +ve
Inhalation of dry yeast forms/basidiospores
Latent lung infection

HIV, TR, IDS
• Meningitis.
• Pneumonia.
• Disseminated infection.
Diagnosis :
CSF : Negative stain.
Tissues : Mayer’s mucicarmine stain, Alcian blue stain.
Culture :
• SDA : Mucoid colonies.
Urease test +ve
• Niger seed medium : Brown colonies.
Capsular antigen detection : Latex agglutination/immunochromatographic test
(ICT)/NAAT in blood samples.
Rx :
No CNS involvement : Fluconazole.
CNS involvement : Amphotericin B + Flucytosine f/b Fluconazole.
Niger seed agar Mucicarmine Negative stain

Dimorphic fungi : Thermal dimorphism 00:50:44 ----- Active space -----
Diseases caused by the fungi are :

Spore tailed Cocks Blasted Para & Histo for being Dimorphic.
• Sporotrichosis.
• Talaromyces.
• Coccidioidomycosis.
• Blastomycosis.
• Paracoccidioidomycosis.
• Histoplasmosis.
All are endemic mycoses except sporotrichosis (India : Sub-Himalayan belt).
All diseases are acquired by inhalation except sporotrichosis (due to trauma).
Sporothrix schenkii :
Aka Rose gardener’s disease.

Symptoms : Nodulo ulcerative lesions along lymphatics, pneumonia (in COPD).
Tissue biopsy : Granulomas with cigar shaped yeasts & asteroid bodies.
Culture at 250C : Hyaline septate hyphae with tear shaped conidia in rosettes.
Sporotrichoid lesions Splendor hoeppli Mold of Sporothrix schenkii

phenomonenon/asteroid
bodies
Rx :
DOC : Itraconazole.
Alternative drugs : Saturated solution of potassium iodide (SSKI), terbinafine.
CNS/disseminated infection : Amphotericin B.

130 10 Microbiology

Sporotrichoid lesions :
Seen in :
• Nodular lymphangitis.
• Lymphocutaneous syndrome.
• Botryomycosis.
• Tularaemia.
• Leishmaniasis.
• Nocardia.
• M. marinum.
H. capsulatum var. capsulatum & var. duboisii :
M/C endemic mycoses.

H. capsulatum var. duboisii is known as African histoplasmosis.
Aka Reticulo endotheliosis/Darlings/ Ohio/Cave’s disease (soil with bat/ bird
droppings).
Symptoms : Acute pneumonitis, mediastinal LN +, chronic cavitary pulmonary
disease in smokers, disseminated in immunodeficiency states
(Lymphadenopathy, hepatosplenomegaly, skin lesions).
Tissue biopsy : Granulomas with budding yeasts in macrophages.
Culture at 250C : Hyaline septate hyphae with microconidia & tuberculate
Macroconidia.
PAS stain GMS stain : Yeast form Hyaline septate

of histoplasma hyphae with
microconidia & tuberculate
Rx : Macroconidia
Mild/moderate : Itraconazole.
Severe : Amphotericin B f/b Itraconazole.
Blastomyces dermatitidis :
Aka Gilchrist/Chicago disease.

Symptoms : Pulmonary, skin papules & nodules, lytic bone lesions, prostatitis,
hepatosplenomegaly, LN +.
Tissue biopsy : Granuloma with broad based ----- Active space -----
budding yeasts showing figure of 8 appearance.
Culture at 250C : Hyaline septate hyphae with
conidia.
Rx :
Mild/moderate : Itraconazole.
Severe : Amphotericin B f/b Itraconazole. Differential interference
contrast microscopy of yeast
Coccidioides immitis/C. posadasii :
Aka Valley fever, San Joaquin disease, desert rheumatism.

Symptoms : Pulmonary, nodular skin lesions, lesions in the skull, spine, swollen
joints, meningitis.
Tissue biopsy : Granulomas with spherules filled by endospores.
Culture at 250C : Hyaline septate hyphae with barrel shaped arthroconidia/
arthrospores.
Barrel shaped anthroconidia Yeast form of

coccidiodes
Rx :
Severe disease : Amphotericin B.
Ketoconazole/itraconazole/fluconazole for 6-12 months.
Paracoccidioides brasiliensis :
Aka South American blastomycosis, Lutz mycosis.

Symptoms : Pneumonia, nasal & oral mucosal lesions,
LN, skin lesions.
Tissue biopsy : Granulomas with yeasts with multiple
Mickey mouse/pilot wheel/
buds (Mickey mouse/pilot wheel/mariner wheel/ mariner wheel/captain wheel
captain wheel appearance). appearance
Culture at 250C : Hyaline septate hyphae with conidia.
Rx :
Amphotericin B, itraconazole, cotrimoxazole.

132 10 Microbiology

Talaromyces (Penicillium) marneffei :
Animal reservoir : Chinese/bamboo rats.

Symptoms : Pneumonia, LN, skin lesions.
Tissue biopsy : Granulomas with ellipsoidal yeasts
dividing by transverse septae.
Culture at 250C : Hyaline septate hyphae with brush
like conidiophore. Red diffusible pigment +.
Red diffusible pigment
Rx :
Amphotericin B followed by itraconazole.
Brush like conidiophore Ellipsoidal yeast dividing

by transverse septae

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Microbiology Revision 1 01 1

MICROBIOLOGY REVISION 1 ----- Active space -----

Stains Used for

Silver impregnation stain Warthin starry stain Negative stain showing

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

----- Active space ----- Steps of gram stain :

Steps of acid-fast stain :

20% sulphuric acid or 3% alchohol (Decolorizer)

AFB : Retain primary stain.

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

Acid-fast structures Partially acid-fast structures

Anatomy of Bacteria 00:10:13

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

----- Active space -----

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

Modes of actions of toxins : ----- Active space -----

Amphitrichous : Campylobacter, Spirillum

Peritrichous : Enterobacteriaceae, Clostridium, Listeria etc

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

----- Active space -----

• Non bulging. • Bulging.

Cultivation of Bacteria 00:30:54

Types of culture medium :

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

agent (To inhibit unwanted bacteria) :

4. Differential medium : Basal/enriched medium + Differentiating agent.

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

α/ β/ γ hemolytic bacteria LF (pink) NLF

Classification of Bacteria 00:42:22

Based on gas requirements :

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

Biochemical Reactions 00:46:00 ----- Active space -----

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

----- Active space ----- Antibiotic Sensitivity Tests 00:50:03

1. Dilution method : 2. Disc diffusion method : 3. Epsilometer/E-test :

Standardisations in antibiotic sensitivity :

Test strain Control strain

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

Intrinsic Resistance of Bacteria 00:54:41 ----- Active space -----

Results from genetic, structural or physiological state.

Bacterial Growth Curve 00:58:05

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

----- Active space ----- • Replicating bacteria = Dying bacteria.

Bacterial growth curve

Bacterial Gene Transfer 01:01:12

Transformation Transduction Conjugation

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

MICROBIOLOGY REVISION 2 ----- Active space -----

Catalase negative bacteria : Catalase Test

Oxidase negative bacteria : Oxidase test (Detection of

Gram positive bacteria

G+ve cocci G+ve bacilli

Staphylococcus Streptococcus Gram Not gram

Gram Positive Cocci 00:02:08

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

----- Active space -----

Slide coagulase test : Tube coagulase test :

β hemolytic, big colonies Golden yellow pigment of S. Yellow colonies of S. aureus

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

Staphylococcus aureus 00:06:27 ----- Active space -----

Slide & tube coagulase +ve.

Microbiology Revision • v1.0 • Marrow 6.5 • 2023

----- Active space -----

2. Exfoliative (Epidermolytic) toxins :