 Bacterial Identification Key

1. Gram Staining:
1. POSITIVE..……………..…………………………………….2
2. NEGATIVE…………………………………………………..19

2. Cell Shape:
1. COCCUS……………………………………………………...3
2. BACILLUS……………………………………………………6

3. Cell Arrangement:
1. STAPHYLOCOCCUS………………………………………..4
2. STREPTOCOCCUS………………………………………......5

4.  Staphylococcus aureus:

5. Streptococcus pyogenes

6. Acid Fast Staining - Ziehl Neelsen:

1. POSITIVE…………………………………………………….7
2. NEGATIVE…………………………………………………...8

7.  Mycobacterium tuberculosis

8. Green Malachite – Spore Staining:

1. POSITIVE…………………………………………………….9
2. NEGATIVE…………………………………………………..14

9. Cell Arrangement or Liquid Medium Turbidity:

1. DIPLO/STREPTOBACILLUS or LOW TURBIDITY……....10
2. ISOLATED CELLS or HIGH TURBIDITY…..……………..13

10. Solid Medium Culture Pigmentation:

1. GREEN PIGMENTATION…………………………………..11
2. NON PIGMENTED…………………………………………..12

11. Bacillus cereus

12. Bacillus anthracis

13. Clostridium perfringens

14.Catalase Test:
1. POSITIVE……………………………………………………..15
2. NEGATIVE……………………………………………………16

15.  Listeria monocytogenes

16.Haemolysis on Blood Agar:

1. BETA and 24HRS……………………………………………...17
2. ALPHA and 48HRS……………………………………………18

17. Arcanobacterium pyogenes

18. Erysipelothrix rhusiopathiae

19.Cell Morphology:
1. BACILLUS……………………………………………………20
2. SPIROCHETE…………………………………………...……37

20.Oxidase Test:
1. POSITIVE……………………………………………………..21
2. NEGATIVE……………………………………………………30

21. Citrate Test:

1. POSITIVE……………………………………………………..22
2. NEGATIVE……………………………………………………27

22. Urease Test:

1. POSITIVE……………………………………………………..23
2. NEGATIVE……………………………………………………24

23. Bordetella sp.

24. Lactose or Indol Test:

1. POSITIVE……………………………………………………..25
2. NEGATIVE……………………………………………………26

25.Aeromonas salmonicida
 26.Pseudomonas aeuriginosaesch

27. Non Haemolytic or Indol Test:

1. POSITIVE……………………………………………………..28
2. NEGATIVE……………………………………………………29

28.Pasteurella multocida:

29.Mannheimia haemolytica

30. Citrate Test:

1. POSITIVE……………………………………………………..31
2. NEGATIVE……………………………………………………36

31. Urease Test:

1. POSITIVE……………………………………………………..32
2. NEGATIVE……………………………………………………35

32. Lactose or H2S Test:

1. LACTOSE POSITIVE / H2S NEGATIVE…………………....33
2. H2S POSITIVE / LACTOSE NEGATIVE….………………...34

33.Klebsiella pneumoniae

34.Proteus mirabillis

35. Salmonella enterica.

36. Escherichia coli

37. Borrelia burgdorfei

Leptospira icterohaemorrhagiae

Gram Negative Biochemical Profile

 Haem
Gram o Cat Ox Cit Ure Lac H2S Ind Mot
Bordetella - a + + + + / - - -
Aeromonas - a + + + - + + - -
Pseudomona
s - b + + + - - - - +
Pasteurella - + + - - - - + -
Mannheimia - a + + - - - - -
Leptospira - / + + / / / / / +
Klebsiella - + - + + + - - -
Proteus - + - + + - + - +
Salmonella - + - + - - + - +
Escherichia - a + - - - + - + +
Borrelia - / / / / / / / / + Ge
neral
Description of Bacteria

 Staphylococcus aureus
-Gram +, small cocci (1 μm), staphylo arrangement, grown on non-enriched
media, Solid: (medium size, white or golden, opaque), Liquid: (medium to
intense turbidity, surface ring), haemolysis dependant on species (aureus alpha),
facultative anaerobe, non-motile
-Catalase +, Coagulase + (correlated with pathogenicity)
-Stable in the environment, cause pyogenic infections (exudative epidermitis in
piglets, tick pyaemia in lambs)

 Streptococcus pyogenes
-Gram +, small cocci (1 μm), strepto arrangement, grown on enriched media,
Solid: (small, circular, translucent, mucoid, 48hrs), Liquid: (low to medium
turbidity, granular/flocculent sediment) usually haemolysis dependant on
species (pyogenes beta)), facultative anaerobe, usually non-motile
-Catalase -
-Commensals of mucous membranes, cause pyogenic infections (Scarlet fever,
septic sore throat, rheumatic fever in humans)

 Listeria monocytogenes
-Gram +, small bacilli (2 μm), grown on non-enriched media, tolerate wide
temperatures and pH, Solid: (small, smooth, flat, blue-green, transparent),
Liquid (low to medium turbidity, cigarette smoke like sediment), beta
haemolysis (rough colonies), facultative anaerobes, mobile at 25C,
-Catalase +, Oxidase -, Camp test + to S. aureus
-Environmental saprophytes, outbreaks often due to spoiled silage feeding, can
lead to encephalitis, abortion, septicaemia or endopthalmitis

 Erysipelotrhix rhusiopathiae
-Gram +, small bacilli (smooth form - 0.4 x 2.5 μm) or filaments (rough form),
grow on non-enriched media, Liquid: (low turbidity, cigarette smoke like
sediment), Solid: (small/pin-point, convex, circular, opaque with irregular
edges), alpha haemolysis after 48 hours
-Catalase-, Coagulase +, H2S +
-Swine erysipelas can be acute: septicaemic or cutaneous (diamond lesions are
pathognomonic) and chronic: vegetative endocarditis and arthritis

 Bacillus anthracis
-Gram +, large bacilli (10 μm), sporulating and capsules (in vivo – Giemsa
staining), arranged in isolated cells or strepto, Liquid: (low turbidity,
granular/flocculent sediment), Solid: (medium to large size, circular or irregular,
opaque, flat, dry, greyish, ground glass after 48hrs, low magnification medusa
head curled colony edges), Ascoli test (thermoprecipitation – antigens), non-
haemolytic, motile
-
-oedema in pigs and horses, intestinal lesions, pulmonary and cutaneous. High
mortality in acute anthrax.

 Bacillus cereus
-Similar to B. athracis but colonies are slightly larger with a greenish colour,
beta haemolysis, non-motile, resistant to penicillin, strong and rapid lecithinase

 Trueperella pyogenes
-Gram +, corynefrom bacilli, arranged in sharp angles-chinese letters-palisades,
Solid: (small/pin-point at 48hrs, convex, smooth, transluscent, circular), Liquid:
(medium turbidity, sediment like cigarette smoke), beta haemolysis,
-
-Mastitis, pneumonia, septic arthritis, vegetative endocarditis, endometritis,
peritonitis, lymphadenitis, omphalitis, seminal vesiculitis, commonly associated
with Fusobacterium. Especially affects cattle, sheep and pigs.
  Clostridium perfringens
-Gram +, short but fat bacilli (0.8-4 μm), endospores (oval and subterminal),
aerotolerant, motile (not C. perfringens), enriched media, Solid: (small to
medium size, irregular, flat, grey/yellowish, opaque or translucent, smooth or
rough), Liquid: (medium to high turbidity, granular/flocculent sediment), double
haemolysis, buthyric smell
-Catalase +, Oxidase –, Camp test + to S. agalactiae, Lactose +, Indole -,
Lecithinase +
-Saprophytes and normal bio-flora, pathogenic effects of their endotoxins
(neurologic, histologic, enterotoxemic- C. perfringens dependent upon the
species)

 Mycobacterium tuberculosis
-Gram +*, acid fast – Ziehl Neelsen staining + or acid/alcohol resistant, slender
bacilli (4 μm long), complex egg enriched media, aerobic, slow growing (3
weeks), Solid: (rough, raised, difficult to break apart, medium to large size,
circular), resistant to chemical disinfectants and the environment, glycerol and
sodium pyruvate present in pathogenic species, tuberculin test for diagnosis
(30+ days post infection)
-Urease +
-Some are saprophytes, others opportunistic or even obligate pathogens
(intracellular). Cause granulomatous lesions (tuberculosis, other species can
produce leprosy or paratuberculosis)

 Escherichia coli
-Gram -, medium sized bacilli, Solid: (moderate, circular, convex, smooth,
characteristic faeces like smell), Liquid: (high turbidity, ring surface structure,
granular or cigarette smoke like sediment), faeces like smell, haemolytic or not,
MacConkey pink
-Lysine +, Glucose +
-Enteric colibacillosis in calves, piglets and lambs. Colisepticaemia in calves,
lambs and poultry. Coliform mastitis.

 Klebsiella pneumoniae
- Gram -, medium sized bacilli, Solid: (mucoid, moderate to large, circular,
uniform, convex, opaque), Liquid: (high turbidity, ring surface structure,
granular or cigarette smoke like sediment
- Lysine +
- Opportunistic pathogen leading to coliform mastitis, endometiris in mares,
pneumonia in calves and foals, UTI in dogs
  Salmonella enterica
-Gram -, medium sized bacilli, peritrichous flagella, Solid: (small to moderate,
circular, convex, shiny), Liquid: (medium to high turbidity, cigarette like smoke
sediment), non-haemolytic, XLD (+ red alkaline with/out black dot H2S if
pathogenic), enriched medium, on brilliant green agar (medium sized and red
colonies), faeces like smell
-Lysine +, VP +, Glucose +
-Enterocolitis, septicaemia, food poisoning, abortion (S. bradenburg)

 Proteus mirabillis
-Gram -, medium sized bacilli, Solid: (swarming growth), Liquid: (high
turbidity, flocculent or cigarette like smoke sediment), faeces like smell
-Lysine –
-Opportunistic bacteria, UTI in dogs and horses, associated in otitis externa,

 Pasteurella multocida
-Gram -, small bacilli (0.2 x 1.5 μm), encapsulated bipolar staining in
Giemsa, Solid: (round, shiny, mucoid in some pathogenic species, sweet odour),
non haemolytic,
-Pneumonia, mastitis, rhinitis, rare cases of peritonitis, septicaemia

 Mannheimia haemolytica
-Similar to Pasteurella sp. But haemolytic, not encapsulated, odourless

 Pseudomonas aeruginosa
-Gram -, medium sized bacilli (1 x 5 μm), obligate aerobe, polar flagella, green-
blue diffusible pigment, Solid: (medium to large sized, irregular, serrate edges,
convex, translucent, shiny or smooth), Liquid: (medium turbidity, ring/pellicle
on the surface, granular/flocculent sediment), grape like odour
-Cause opportunistic infections (mastitis, pneumonia, dermatitis, otitis media,
genital tract infections, cystitis, and septicaemia)

 Aeromonas salmonicida
-Gram -, medium sized (0.5 x 3 μm) bacilli straight or curved, grow on non-
enriched media, facultative anaerobes, polar flagella, often arranged in pairs-
chains-clusters, Solid: (small to medium size, circular, uniform, convex,
translucent, smooth), Liquid: (high turbidity), haemolysis after 48hrs, brown
pigment on Difco agar.
-Aquatic environments, opportunistic pathogens mainly of fish and reptiles
  Bordetella sp.
-Gram -, small bacilli (0.4 x 1 μm), peritrichous flagella, grow on non-enriched
media, majority of pathogenic species grow on MacConkey (lactose dependent
on species), strict aerobes, toxigenic strains agglutinate mammalian RBC, Solid:
(convex, small, smooth, whitish, circular), haemolytic or not species dependant
-Commensals in upper respiratory tract can lead to opportunistic infections
(pneumonia, rhinitis, tracheobronchitis)

 Leptospira icterohaemorrhagie
-Gram -, spirochete with endoflagella (0.1 x 12 μm), motile, helical, some only
grow on liquid media, most require specialized media, dark field microscopy,
silver staining, some slow growing serotypes such as hardjo can take up to 6
months
-Labile in the environment (very sensitive to dessication), many produce
zoonotic diseases, systemic infections, contaminated urine

 Borrelia burgdorfei
-Gram X, spirochete (8-20 μm), motile, helical, slow growing, microaerophilic,
Barbour-Stoenner-kelly medium (6 weeks)
-Systemic infections in many species, transmitted by arthropod vectors (ticks),
known for Lyme disease and avin borreliosis.