PSEUDOMONAS

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PSEUDOMONAS

• Introduction

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• Large group of aerobic
• non sporing
• gram negative
• motile by polar
  flagella
• ubiquitous
• oppurtunistic infections
• newer genera-Burkholderia
• Stenotrophomonas

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Species

• Pseuomonas aeruginosa
• Stenotrophomonas maltophila
• Burkholderia cepacia
• Burkholderia mallei
• Burkholderia pseudomallei

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Pseudomonas aeruginosa

• Morphology
• slender,Gram negative bacillus
• size-1.5 microns-31.5microns
• motile by polar flagella
• non capsulated though some mucoid strains may
  sometimes occur
• some are pilated

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Cultural characteristics

• Obligate aerobe
• Wide range of temperature 5c-42c
• optimum 37c
• Ordinary media
• large,opaque,irregular,with distinctive
  musty,mawkish,earthy smell

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Cont..

• Nutrient agar-
• Colonies are smooth,large,translucent,low
  convex,2-4mm in diameter.
• Produce sweetish aromatic odour
• Greenish blue pigment diffuses

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Cont..

• Blood agar
• Similar to nutrient agar
• Many are haemolytic
• Mac conkey agar
• Colourless,non lactose fermenters
• Cetrimide agar
• selective media

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Pigment production

• Pyocyanin
• Bluish green phenazine pigment
• Soluble in chloroform and water
• Not produced by other species
• Pyoverdin(fluorescin)
• It is a greenish yellow pigment
• Insoluble in chloroform but soluble in water
• Produced by many other species

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Pyocyanin Pyoverdin

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Cont..

• Pyorubin
• Reddish brown pigment
• Insoluble in chloroform but soluble in water
• pyomelanin
• Brown to black pigment
• Production is uncommon

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Biochemical reactions

• O/F test-oxidative
• Catalase-positive
• Oxidase-positive
• Nitrate reduction-positive

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Cont..

• Indole test-negative
• Methyl red test-negative
• Vp test-negative
• Citrate test-positive
• Urease test-negative

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Cont

• Sugar fermentation tests
• Glucose-only acid
• Lactose-negative
• Sucrose-negative
• Mannitol-negative

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Resistance

• Susceptible to
• heat,killed at 55c in one hour
• acids
• beta glutarldehyde
• Silver salts
• strong phenolic disinfectants
• So,silver sulphonamide as a topical cream in
  burns is used

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Cont

• Resistant to
• chemical agents
• common antiseptics
• disinfectants -ammonium compounds
  chlroxylenol
• hexachlorophane
• antibiotics -aminoglycosides
• cephalosporins
• pencillins

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Antigenic structure

• O antigens
• Possesses 19 distinct,group specific O antigens
• Heat labile
• H antigens
• Two heat labile antigens recognised in
  Ps.aeruginosa

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Toxins and enzymes

• Extracellular products
• Pyocyanin inhibits mitochondrial enzymes and
  causes disruption and cessation of ciliary beat
  on ciliated nasal epthelium
• Extracellular enzymes and haemolysins
• Produces proteases,haemolysins and lipases
• Exotoxins
• Produces two exotoxns A and S
• Endotoxins
• It is a lipopolysaccharide have pyrogenic
  action

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Typing methods

• Bacteriocin typing
• Three types of bacteriocins are produced-R,F,S
• Pyocin produced by test strain is employed to
  assess the growth inhibition of 13
• Depending upon the growth inhibition of 13
  indicators strains,105 types are recognised
• Most popular method used

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Cont..

• Phage typing
• Serotyping
• based on O and H,17 serotypes of
  Ps.aeruginosa are recognised
• Molecular Method
• Restriction endonuclease typing with
  pulsed-field gel electrophoresis(PAGE) is most
  reliable

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PATHOGENESIS

• Important agent in causing nosocomial infections
• Most common infections are
• Urinary tract infections following
  catheterisation
• Acute purulent meningitis following lumbar
  puncture
• Post-tracheostomy pulmonary infection
• Septicaemia in debilitated patients

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cont..

• Wound and burn infections
• Chronic otitis media and otitis externa
• Eye infections
• Acute necrotising vasculitis
• Infantile diarrhoea

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Mechanism Of Pathogenesis

• Caused by exotoxins,proteases,elastases,hae
  molysins,lipases and enterotoxins
• Exotoxin A-lethal toxin
• Elastases-haemorrhagic lesions
• Enterotoxins-diarrhoeal disease
• Slime layer acts as a capsule and enhances
  virulence

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• Thank you

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HAEMOPHILUS

• INTRODUCTION

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HAEMOPHILUS INFLUENZAE

• MORPHOLOGY
• SMALL 1.0 X 0.3 MICRONS
• GRAM NEGATIVE
• NON MOTILE
• NON SPORING
• EXHIBIT PLEOMORPHISM

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CULTURAL CHARACTERISTICS

• FACTOR X

• FACTOR V

• HEAT STABLE
• HEMIN
• AEROBIC RESPIRATION

• HEAT LABILE
• BACTERIAL VITAMIN
• ANEROBIC RESPIRATION

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CONT

• AEROBIC
• GROWS ANEROBICALLY ALSO
• OPTIMUM TEMPERATURE-37C
• SOME STRAINS REQUIRE 10 CO2
• BLOOD AGAR-SCANTY GROWTH

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SATELLITISM

• Staph.aureus is streaked across a plate of blood
  agar on which a specimen containing H.influenzae
  has inoculated
• After overnight incubation-
• Colonies
• large -along the streak of Staph.aureus
• small farther away

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satellitism
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CONT

• LEVINTHALS MEDIUM
• Boiling and filtering a mixture of blood and
  nutrient broth
• Capsulated strains produce translucent colonies
• FILDES AGAR
• By adding a peptic digest of blood to nutrient
  agar
• Best for primary isolation
• Copious growth

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BIOCHEMICAL REACTIONS

• Catalase-positive
• Oxidase-positive
• Glucose,Xylose-fermented with acid production
• Lactose,Sucrose,Mannitol-not fermented

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RESISTANCE

• Destroyed by heating
• refrigeration
• drying
• disinfectants
• Cultures may be preserved for about a month on
  choclate agar slopes in screw capped bottles
• Long term preservation culture may be
  lyophilised

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ANTIGENIC PROPERTIES

• 3 Major surface antigens
• Capsular polysaccharide
• Outer membrane protien
• Lipo oligosaccharide
• Major antigenic determinant of capsulated strains
  Capsular polysaccharide
• Pittman classified into 6 capsular strains i.e
  type a to f

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• Meningitis belong to type b strain
• Type b strain contains pentose sugars ribose
  ribitol instead of hexoses hexosamines in
  other 5 serotypes

• Capsular polyribosyl ribitol phosphate antigen of
  Hib induces IgG IgM IgA antibodies which are
  bactericidal and protective

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CONT.

• Strains lacking a capsule cannot be typed and are
  called NON TYPABLE STRAINS
• Outer membrane protein antigens of H influenzae
  type b have been classified into 13 sub types
• Lipo oligosaccharides are antigenically complex

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PATHOGENECITY

• Exclusively human pathogen
• Not pathogenic for animals but intra peritoneal
  inoculation of large doses is fatal in
  mice,guinea pigs rabbits
• Diseases due to H influnzae considered under 2
  groups invasive and non invasive

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INVASIVE Spread through Blood Meningitis,arthritis, endocardits,conjuctivitis,pancarditis Seen in children Caused by capsulated strains NON INVASIVE Spread through local invasion Otitis media,sinusitis Seen in adults Caused by non capsulated strains

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CLINICAL MANIFESTATIONS

• MENINGITIS
• Bacilli reach meninges from nasopharynx through
  blood stream
• Common in children
• LARYNGO EPIGLOTTIS
• Acute inflammaton of epiglottis with obstructive
  laryngitis
• Children above 2 years

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CONT.

• PNEUMONIA
• Infants accompanied by empyema
• Older children adults by lobar pneumonia
• These are primary infections due to capsulated
  strains
• Broncho pneumonia may occur as secondary
  infection due to non capsulated strains

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CONT

• SUPPURATIVE LESIONS
• Arthritis,endocarditis pericarditis may result
  from hematogenous dissemination
• Otitis media occurs by direct spread from
  nasopharynx
• BRONCHITIS
• Associated with pneumococci in acute
  exacerbationof chronic bronchitis bronchiectasis

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LAB DIAGNOSIS

• 1.SPECIMENS
• CSF
• Blood
• Throat swab
• Sputum
• Pus
• Aspirates from joints ,middle ears

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2.COLLECTION TRANSPORT

• Collected in sterile containers under aseptic
  conditions
• Very sensitive to low temp so should not be
  refrigerated
• Transported to laboratory with out delay
• Inoculated on culture media immediately

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CONT.....

• 3.DIRECT MICROSCOPY
• Gram staining
• Immunoflouroscence and quellung reaction
• Antigen detection
• Latex agglutination
• Co-agglutination
• Counter immuno electrophoresis

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CONT

• 4.CULTURE
• CSF culture
• Blood culture
• Sputum culture
• 5.COLONY MORPHOLOGY AND STAINING
• 6.SEROTYPING

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TREATMENT

• Cefotaxime Ceftazidime is the drug of choice
  for meningitis
• Ampicillin Cotrimoxazole respiratory infections
• Amoxycillin - clavulanate or clarithromycin is
  more effective

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EPIDEMIOLOGY PREVENTION

• Infection transmitted by respiratory route
• Immunity is type specific
• Active immunization with Hib PRP vaccine is
  indicated
• Conjugate Hib PRP is used for younger children
• Rifampicin -4 days prevents secondary infection
  in contacts also eradicates carrier state

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Haemophilus aegyptius

• KOCH WEEKS BACILLUS
• Non capsulated
• Causes highly contagious form of conjuctivitis
  pink eye
• Causative agent of Brazilian purpuric fever BPF
  
• Responds to local sulphanamides gentamicin

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Haemophilus ducreyi

• Ducrey demonstrated in chancroid lesions
• Short ,ovoid bacillus
• Gram nagative often may appear gram positive
  frequently show bipolar staining
• May be arranged in small groups or whorls or in
  parallel chains giving a SCHOOL OF FISH or
• RAIL ROAD TRACK appearance

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CONT

• Can be grown on fresh clotted rabbit blood
• May also grown on chorio allontioc membrane of
  chick embryo
• On choclate agar enriched with isovitalex and
  fetal calf serum containing vancomycin as a
  selective agent
• Forms small ,grey ,translucent colonies after
  incubation at 35 c under 10 co2 high humidity
  in 2 8 days

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CONT

• Antigenically homogenous
• Cultures may be identified by agglutinatoin with
  anti serum
• Susceptible to sulphonamides many antibiotics
• Erythromycin ,Cotrimoxazole ,Ciprofloxacin may be
  used for treatment

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Choclate agar
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Haemophilus parainfluenzae

• Requires factor V not the factor X
• Commensel in the upper respiratory tract
• Causes bacterial endocarditis ,urethritis ,acute
  pharyngitis

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Haemophilus haemolyticus

• Requires both factor X V
• Commensal of upper respiratory tract
• Strains that donot require factor X have been
  designated as H parahaemolyticus

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Haemophilus aphrophilus

• Requires factor X but not factor V
• Cause bacterial endocarditis ,brain abscess
  ,sinuisitis pneumonia
• Strains requiring factor V but not factor X have
  been termed as H paraaphrophilus

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HACEK group bacteria

• Fastidious slow growing bacteria
• H - Haemophilus species
• A - Actinobacillus
  C - Cardiobacterium hominis
• E - Eikenella corrodens
• K - Kingella kingae

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• Thank you