Thursday, August 30, 2012

Listeria sp. Identification Test

Listeria sp.


Morphology: Rods
Gram: Positive
Catalase: Positive
Respiratory system: Facultative anaerobe 
Motility: Peritrichous

Tests to identify the genus

  • Hemolysis test - Sheep blood agar
  • CAMP factor - Sheep blood agar
  • Mixed acid or butane-diol fermentation - Clark and Lubs medium
  • Sugar fermentation: xylose, mannitol, rhamnose
  • Nitrate reductase

Culture

PALCAM agar

Listeria sp. on PALCAM agar
On BD PALCAM Listeria Agar, colonies of Listeria appear gray-green with a black halo. Confirmation of the presence of Listeria is made following subculture onto appropriate media and biochemical/serological identification. Colonies of mannitol-fermenting organisms such as staphylococci, which may grow on this medium, appear yellow with a yellow halo.

Refererence: BD™ [Link]







ALOA™ agar
Listeria monocytogenes on ALOA™












Please, feel free to comment if there is any mistake.

Bacillus sp. Identification Test

Bacillus sp.


Morphologie: Rods
Gram: Positive
Catalase: Positive
Respiratory system: Facultative aerobe (B. subtilis: Obligate aerobe)
Motility: Peritrichous
Characteristic: Produces endospores

Tests to identify spieces

  • CO2 production - Gibson Abdel Malek (GAM) medium
  • Acetyl-methyl-carbinol (AMC) production - AMC medium
  • Indole production - Peptone water indole-free
  • Urease test - Christensen medium
  • Citrate utilization - Simmons medium
  • Growth at 65° C
  • 7 % NaCl resistance
  • Proteolysis test
  • Gelatinase test
  • Amylase test
  • Hemolysis - Sheep blood agar
  • Sugar fermentations: glucose, arabinose, mannitol and xylose
Please, feel free to comment if there is any mistake.

Clostridium sp. Identification Test

Clostridium sp.


Morphology: Rods
Gram: Positive
Catalase: Negative
Oxidase: Negative
Respiratory system: Obligate anaerobe
Motility: Motile
Characteristic: Produces endospores

Tests to identify the genus

  • API 20A
  • Rosenow medium
  • Nitrate reductase test
  • Proteolysis and acidification
  • Sulfites reduction
  • Lecithinase test

Test to identify butyric Clostridium

  • Lactate utilization: butyric acid and CO2 production - Bryant and Burkey medium
Please, feel free to comment if there is any mistake

Lactic Acid Bacteria (rods) Identification Test

Lactic acid bacteria (rods)

Lactobacillus casei, L. plantarum, L. fermentum, L. acidophilus, L. bulgaricus and L. helveticus

Morphology: Rods
Gram: Positive
Catalase: Negative
Motility: Non-motile
Respiratory system: Facultative aerobe



Tests to identify the genus Lactobacillus 

  • CO2 production from glucose
  • Culture at 15° C and 45° C - MRS medium
  • Arginine dihydrolase - Moeller medium
  • Sugar fermentation : ribose, lacotse, maltose, sorbitol or sucrose

Test to identify the species

  • Esculin hydrolysis - Esculin agar
  • Acidification and reduction

Please, feel free to comment if there is any mistake.

Lactic Acid Bacteria (cocci) Identification Test

Lactic acid bacteria

Lactococcus lactis, Lactococcus lactis subsp. lactis biovar diacetylactis, Lactococcus lactis subsp. cremoris, Streptococcus thermophilus, Pedicoccus acidilactici and Leuconostoc sp.

Morphology: Cocci
Gram: Positive
Catalase: Negative
Respiratory system: Facultative anaerobe


Tests to identify the genus

  • Growth at 10° C and 45° C - Elliker/M17 broth
  • NaCl resistance: 2, 4, 6 % NaCl
  • CO2 production - MRS medium with bell

Tests to identify the species

  • Acidification and reduction test
  • Citratase test
  • Arginine dihydrolase test - Moeller medium
  • Sugar fermentations: glucose, sucrose, ribose, xylose, maltose, lactose
Please, feel free to comment if there is any mistake.

Fecal contamination indicators

Fecal contamination indicators

Enterococcus fæcalis (Streptococcus fæcalis), Enterococcus fæcium (Streptococcus fæcium), Enterococcus durans (Streptococcus durans) and Streptococcus equinus (Streptococcus bovis)


Morphology: Cocci
Gram: Positive
Catalase: Negative
Respiratory system: Facultative anaerobe
Characteristic: bile-resistant


Characteristics

  • Growth at 10° C and 45° C
  • Temperature resistance - 60° C for 30 minutes
  • Methylene blue reduction - Sherman's milk
  • NaCl-resistant: 6,5 %

Identification tests

  • Tellurite resistance - Potassium tellurite agar
  • Tetrazolium chloride (TTC) reduction - Barnes medium
  • Hemolysis - Sheep blood agar (5 %)
  • Gelatinase - gelatine broth
  • Sugar fermentation (lactose, sucrose, mannitol, arabinose, sorbitol)

Culture

Slanetz & Bartley medium

Selective enumeration of Enterococci in water
Sodium azide inhibits the growth of Gram-negative bacteria. TTC is an indicator of bacterial growth. It is reduced to an insoluble formazan inside the cells. This reaction is seen by the formation of red to maroon colonies.

Retain only membranes containing fewer than 100 colonies. Red, maroon or pink colonies are 
considered as characteristic. Carry out confirmation of typical colonies using Bile Esculin Azide Agar.

Reference: Biokar Diagnostics [Link]

Bile Esculin Agar

Organisms positive for esculin hydrolysis hydrolyze the esculin to esculetin and dextrose. The esculetin reacts with the ferric citrate to form a dark brown or black complex. Oxbile is used to inhibit Gram-positive bacteria other than enterococci. Beef Extract and Enzymatic Digest of Gelatin are the carbon and nitrogen sources used for general growth requirements in Bile Esculin Agar. Agar is the solidifying agent.

Reference: Neogen [Link]

Please, feel free to comment if there is any mistake.


Microccus & Staphylococcus Identification Test

Microccus & Staphylococcus


Morphology: Cocci
Gram: Positive
Catalase: Positive

Differences between Micrococcus and Staphylococcus

CharacteristicsMicrococcus spp.Staphylococcus spp.
Respiratory systemObligate aerobeFacultative anaerobe
OxidasePositiveNegative
Resistance to vibriostatic agent O/129SR
Resistance to lysostaphinRS (±R)
Resistance to bacitracinSR
Growth in presence of furazolidone+-
% GC66-75 %30-39 %


Tests to identify pathogenic Staphylococci

  • Free coagulase test - Coagulase broth
  • Clumping factor - Slidex staph kit
  • Deoxyribonuclease (DNAse) test
  • Thermostable DNAse test - Lachica medium
  • Hemolysis test - Sheep blood agar (5 %) 

Cultures

Baird-Parker Agar


Principles: 
Enzymatic Digest of Casein and Beef Extract are the carbon and nitrogen sources in Baird Parker Agar. Yeast Extract supplies B-complex vitamins that stimulate bacterial growth. Glycine and Sodium Pyruvate stimulate growth of staphylococci. The selectivity of the medium is due to Lithium Chloride and a 1% Potassium Tellurite Solution, suppressing growth of organisms other than staphylococci. The differentiation of coagulase-positive staphylococci is based on Potassium Tellurite and Egg Yolk Emulsion. Staphylococci that contain lecithinase break down the Egg Yolk and cause clear zones around the colonies. An opaque zone of precipitation may form due to lipase activity. Reduction of Potassium Tellurite is a characteristic of coagulasepositive staphylococci, and causes blackening of colonies. Agar is the solidifying agent.

Results:
Staphylococcus aureus on Baird-Parker agar
Coagulase-positive staphylococci produce black, shiny, convex colonies with entire margins and clear zones. Sometimes, an opaque zone of precipitation may form due to lipase activity around that clear zone. Coagulase-negative staphylococci produce poor or no growth. If growth occurs, colonies are black; clear or opaque zones are rare. The majority of other organisms are inhibited or grow poorly. If growth appears, colonies are light to brown-black, with no clear or opaque zones. 

Reference: Neogen [Link]





Baird-Parker RPF (Rabbit Plasma Fibrinogen)

Champan agar


Staphylococcus aureus on Chapman agar













Please, feel free to comment if there is any mistake 

Eterobacteriaceae Identification Test

Enterobacteriaceae


Morphology: Rod-shape
Gram: Negative
Oxidase: Negative
Motility: Peritrichous (except Klebsiella sp. & Shigella sp.)
Respiratory system: Facultative anaerobe

Tests to identify genus & species

Carbon metabolism

  • Methyl red & Voges Proskauer - Clark & Lubs medium
  • Glucose and lactose fermentation, gas and H2S production - Kligler medium
  • β-galactosidase - ONPG
  • Citrate utilization - Simmons medium
  • Malonate utilization - Leifson medium

Nitrogen metabolism
  • Gelatinase test
  • Amino-acids deamination
  • Lysine decarboxylase - Taylor or Moeller media with and without Lysine
  • Indole production - Peptone water indole-free
  • Urease - Christensen medium

IMViC - Indole Methyl red Voges Proskauer Citrate

Micro-organismIndoleMRVPCitrate
E. coli++--
Edwardsiella++--
Proteus sp.++--
Klebsiella pneumoniae-+-+
Klebsiella oxytoca+-++
Enterobacter sp.--++
Serratia sp.--++
Citrobacter freundii-+-+
Citrobacter koseri++-+

Please, feel free to comment if there is any mistake.