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This document provides information on various microbiology tests used to identify bacterial species, including examples of positive and negative results. It describes tests such as the catalase test to distinguish Staphylococci from Streptococci, the coagulase test for identifying Staph aureus, and the DNase test to differentiate Staph aureus from Staph epidermidis. It also summarizes culture-based tests on different media types to isolate and identify bacteria such as Salmonella, E. coli, and Streptococcus species.
Slides examبكتريا عملي
- 1. Diagnostic Microbiology Lab. By Dr. Nabil El Aila Assistant Professor of Molecular Microbiology Medical Technology Department Al -Aqsa University
- 2. A B Name of the test: Catalase test Example A: positive - Staphylococci Example B: Negative - Streptococci Principle: 2 H2O2 2 H2O + O2 Catalase
- 3. A B Name of the test: Coagulase test Example A: Positive - Staph aureus Example B: Negative - Staph epidermedis Principle: Fibrinogen in plasma Fibrin clot Coagulase
- 4. A B Name of the test: DNase test Example A: Positive - Staph aureus Example B: Negative - Staph epidermedis Priciple: DNA is insoluble in acid DNA is hydrolyzed into oligonucleotides by the action of Dnase Nucleotides soluble in acid (After addition of 1N HCl)
- 5. Name of the test: Mannitol salt agar Example A: Negative - Staph epidermidis Example B: Positive - Staph aureus Principle: Staph aureus ferment mannitol (sugar) and phenol red (pH indicators turns yellow in acidic pH and turns red in alkaline pH). A B
- 6. Name of the test: Novobiocin sensitivity Example A: Staph epidermidis (Sensitive) Example B:Staph saprophyticus (Resistant) A B
- 7. Name of the test: Bacitracin sensitivity Example A: Strept pyogens (sensitive) Example B: Strept agalactiae (resistant) A B
- 8. A B Name of the test: CAMP test Example A: Positive - Strept agalactiae (Arrow shaped) Example B: Negative - Strept pyogens Principle : Strept agalactiae produce CAMP factor (a diffusible extracelluar protein) that synergistically acts with the beta-lysin of Staphylococcus aureus and enhances the lysis of red blood cells.
- 9. Name of the test: Optochin sensitivity Example A: Strept pneumonia (sensitive) Example B: Strept viridans (resistant) A B
- 10. A B Name of the test: Bile Esculin test Example A: Positive - Group D streptococcus (Enterococcus species) Example B: Negative - Group B streptococcus Principle: The selective agent bile, inhibits most gram positive bacteria. Esculin in the medium is hydrolyzed to esculetin and dextrose. The esculetin reacts with ferric chloride in the media to form a black-brown color.
- 11. B A Name of the test: Bile Solubility Example A: Strept mitis (Negative) Example B: Strept pneumoniae (positive) Principle: S. pneumoniae is bile soluble whereas all other alpha-hemolytic streptococci are bile resistant. Sodium deoxycholate (2% in water) will lyse the pneumococcal cell wall
- 12. A B Name of the test: Pyrrolidonyl arylamidase Test (PYR) Example A: Strept agalactiae (Negative) Example B: group A streptococci and Enterococcu s species (positive) Principle: Group A streptococci and Enterococcu s species produce pyrrolidonyl arylamidase which hydrolyzes the substrate L- pyrrolidonyl -β- naphthylamide to form β-naphthylamine. A pink to red color forms when p-dimethylaminocinnam-aldehyde (PYR reagent) is added to β-naphthylamine.
- 13. A B Tryptophane amino acids Tryptophanase Indole + Pyurvic acid + NH3 Kovac’s Reagent Red color in upper organic layer` Name of the test: Indole test Example A: Klebsiella pneumonia (Negative) Example B: Escherichia coli (Positive) Indicator: Kovac’s Reagent (4 (p) – dimethylamino benzaldehyde) Principle:
- 14. A B Name of the test: Methyl red Example A: Klebsiella pneumonia (Negative) Example B: Escherichia coli (Positive) Indicator: Methy red indicator Principle: follow next slides
- 15. A B Name of the test: Voges Proskauer test Example A: Escherichia coli (Negative) Example B: Klebsiella pneumonia (Positive) Indicator: -naphthol and 40% KOH Principle: follow next slide
- 16. IMViC test Methyl Red-Voges Proskauer (MR-VP) Tests Glucose Acidic pathway Mixed acids pH less than 4.4 Methyl Red indicator Red color Principle MR positive E. coli Or Neutral pathway Acety methyl carbinol (ACETOIN) Barrit’s A (-naphthol) Barrit;s B (40% KOH) Pink color VP positive Klebsiella
- 17. Name of the test: Citrate test Example A: Klebsiella pneumonia (Positive) Example B: Escherichia coli (Negative) Indicator: Bromothymol blue Principle: follow next slide A B
- 18. Principle: Citrate Na2CO3 Alkaline,↑pH Bromothymol blue Simmone’s Citrate media Pyruvate CO2 + Na + H2O Positive test Contains Citrate as a sole of C source IMViC: CITRATE TEST Dr. Nabil El Aila Diagnostic Microbiology
- 19. Name of the test: Urea test Example A: Proteus miribalis (Positive) Example B: Escherichia coli (Negative) Indicator: Phenol red Principle: A B Urea Urease CO2 + NH3 H2O NH4 OH ↑ in pH Phenol Red Pink Positive test
- 20. A B Name of the test: Motility test Example A: Escherichia coli (Negative) Example B: Proteus miribalis (Positive)
- 21. A B Name of the test: H2S production Example A: Escherichia coli (Negative) Example B : Proteus miribalis (Positive) Principle: Bacteria use enzyme cysteine desulfurase to hydrolyze the amino acid cysteine, forming hydrogen sulfide as end-product. The hydrogen sulfide will combine with the iron salt to form a visible black ferric sulfide (FeS) in the tube
- 22. A B Name of the test: Triple sugar Iron Example A: K/A - No H2S – no gas production Glucose fermenter - Shigella species Example B: A/A - No H2S – gas production All three sugars fermented – Escherichia coli
- 23. A B Name of the test: Triple sugar Iron Example A: A/A - H2S positive –gas production Glucose fermenter - Proteus species Example B: K/A - H2S positive – gas production All three sugars fermented – Salmonella species
- 24. A B Name of the test: Triple sugar Iron Example A: K/K – No H2S – No gas production No action on sugars - Pseudomonas species Example B: K/A – No H2S – No gas production Glucose fermenter - Shigella species
- 25. Name of the test: Lysine Decarboxylase Agar Example A: Positive decarboxylation (butt), negative deamination (slant) : Salmonella species Example B: Negative decarboxylation (butt), Positive deamination (slant) : Proteus species A B
- 26. A B Name of the test: Lysine Decarboxylase Agar Example A: Positive decarboxylation (butt), negative deamination (slant) : Salmonella species Example B: Negative decarboxylation (butt), Negative deamination (slant) : Citrobacter species
- 27. Name of the test: Oxidase test Example A: Positive - Pseudomonas species Example B: Negative - Proteus species Princile: follow next slide A B
- 28. Oxidase Test: Principal Oxidase Reagent Cytochrome Oxidase Indophenol Play role in aerobic respiration Pseudomonas Vibrio Alternative substrate for Cytochrome Tetramethyl-P- Pheneylenediamine Colorless Purple color Oxidize the reagent from colorless to purple color
- 29. A B Name of the test: Oxidation Fermentation test Example A: Oxidation negative (Open tube remain green) Example B: Fermantation negative ( Covered tube remain green) : Oxidation negative/ Fermentation negative (Alcaligenes faecalis)
- 30. A B Name of the test: Oxidation Fermentation test Example A: Oxidation positive (Open tube turns yellow) Example B: Fermantation negative ( Covered tube remain green) : Oxidation positive/ Fermentation negative (Pseudomonas species)
- 31. A B Name of the test: Oxidation Fermentation test Example A: Oxidation positive (Open tube turns yellow) Example B: Fermantation positive( Covered tube turns yellow) : Oxidation positive/ Fermentation positive (Enterobacteriacea)
- 32. Name of the test: Gelatin Liquification test Example A: Negative (Proteus species) Example B: Positive ( Pseudomonas species) Principle : Nutrient gelatin Protein/Polypeptides Solid Gelatinase Incubation at 37/overnight Nutrient gelatin Amino acids Liquid at > 25 C Gelatinase hydrolyze the protein to aminoacids Pseudomonas A B
- 33. A B C D Name of the test: Nitrate reduction test Example A: Negative nitrate reduction - no change with alpha naphthol and sulf.acid Example B: Positive nitrate reduction – turns red with alpha naphthol and sulf.acid sulf.acid Example C: Positive nitrate reduction . No red color after addition of zinc dust ( Nitrate reduced into nitrite and further reduction to Nitrogen. Example D: Negative nitrate reduction red color after addition of zinc dust Nitrate unreduced Principle : follow next slide
- 34. Nitrate Reductase Test: Principal Nitrate (NO3) Nitrate reductase Nitrite (NO2) α-naphthylamine Sulfanilic acid Red diazonium salt If no red color! Further reduction Nitrogen gas N2 Add zinc dust (reducing agent) Nitrate Reductase No red color after addition of zinc dust ( Nitrate reduced into nitrite and further reduction to Nitrogen (Positive nitrate reduction ) or red color after addition of zinc dust Nitrate unreduced (Negative nitrate reduction )
- 35. Name of the test: Satellitism Principle: - A lawn of test bacteria is plated onto a fresh blood agar plate. This provides a supply of haem. - An inoculum of Staphylococcus aureus, which can provide NAD, is placed on the plate and the culture is incubated. - Haemophili can be seen growing larger near to the staphylococcal colony, where the supply of NAD is greatest.
- 36. Name of the test: Swarming Example: Proteus species move very actively by means of peritrichous flagella, resulting in "swarming" on solid medium
- 37. Name of the test: Exopigment production on Muller Hinton agar Example: pseudomonas species produces exopigments in culture eg pyocyanin, fluorescein, pyorubrin, pyoverdin, and pyomelanin
- 38. Bacteria grow in the presence of X factor Example: Haemophillus aphrophilus
- 39. Bacteria grow in the presence of V factor Example: Haemophillus parainfluenza
- 40. A B A: Vaginal saline preperation: normal epithelial cells B: Vaginal saline preperation: Clue cells
- 41. Gram variable bacteria (Gram negative and positive coccobacilli) Gardnerella vaginallis
- 42. Name of test: Motility test listeria A: Non motile B: umbrella shape indicative of motility A B
- 43. A: Dark field microscopy B: Fluorescence microscopy antibody staining Dr. Nabil El Aila Diagnostic Microbiology A B
- 44. Grram positiv cocci in Clusters Example: Staph aureus Dr. Nabil El Aila General Microbiology
- 45. Grram positiv cocci in Tetrads Example: Micrococcus species Dr. Nabil El Aila General Microbiology
- 46. Grram positiv cocci in Chains Example: Streptococcus pyogens Dr. Nabil El Aila General Microbiology
- 47. Grram positiv cocci in pairs Example: Streptococcus pneumoniae Dr. Nabil El Aila General Microbiology
- 48. Gram negative coccobacilli Example: Haemophilus influenza Dr. Nabil El Aila General Microbiology
- 49. Grram positiv bacilli (spore forming) Example: Bacillus anthracis Dr. Nabil El Aila General Microbiology
- 50. Gram positiv bacilli (non spore forming) Example: Lactobacillus species Dr. Nabil El Aila General Microbiology
- 51. Gram positiv bacilli (non spore forming) (Chines letter) Example: Corynebacterium diphtheria Dr. Nabil El Aila General Microbiology
- 52. Gram Negative diplococci Example: Neisseria gonorrhoeae Neisseria meningitidis Dr. Nabil El Aila General Microbiology
- 53. Gram Negative bacilli Example: Escherichia coli Dr. Nabil El Aila General Microbiology
- 54. Gram Negative curved rods (Comma shaped) Example: Vibrio cholera Dr. Nabil El Aila General Microbiology
- 55. Acid fast bacilli Example: Mycobacterium tuberculosis Dr. Nabil El Aila General Microbiology
- 56. Gram Negative spiral rods Example: Campylobacter species Dr. Nabil El Aila General Microbiology
- 57. Spirochates Example: Treponema pallidum
- 58. Type of culture media: Basic Media Example: Nutrient agar Microbe: Staph epidermidis
- 59. Type of culture media: Enriched Media Example: Blood agar (golden yellow) Microbe: Staph aureus Dr. Nabil El Aila General Microbiology
- 60. Type of culture media: Enriched Media Example: Blood agar (white colonies) Microbe: Staph epidermidis Staph saprophyticus Dr. Nabil El Aila General Microbiology
- 61. Type of culture media: Enriched Media Example: Blood agar (Alpha haemolysis) Microbe: Strept pneumoniae
- 62. Type of culture media: Enriched Media Example: Blood agar (Beta haemolysis) Microbe: Strept pyogens
- 63. Type of culture media: Enriched Media Example: Blood agar (Gamma haemolysis) Microbe: Enterococcus faecalis
- 64. Type of culture media: Enriched Media Example: Chocalate agar Microbe: Haemophilus influenza Dr. Nabil El Aila General Microbiology
- 65. Type of culture media: Differential Media Example: MacConkey agar Microbe: Escherichia coli
- 66. Type of culture media: Differential Media Example: MacConkey agar Microbe: Klebsiella pneumoniae
- 67. Type of culture media: Differential Media Example: TCBS (Thiosulfate citrate bile salts sucrose agar) Microbe: Vibrio cholera Dr. Nabil El Aila General Microbiology
- 68. Type of culture media: Differential Media Example: CLED (Cystine-Lactose-Electrolyte-Deficient Agar ) Microbe: Escherichia coli Dr. Nabil El Aila General Microbiology
- 69. Type of culture media: Selective Media Example: Salmonella shigella agar Microbe: Salmonella typhi
- 70. Type of culture media: Selective Media Example: Xylose lysine Deoxycholate Microbe: Salmonella typhi
- 71. Type of culture media: Selective Media Example: Hecktoen Enteric Microbe: Salmonella typhi Dr. Nabil El Aila General Microbiology
- 72. Type of culture media: Selective Media Example: Mannitol salt agar Microbe: Staph aureus Dr. Nabil El Aila General Microbiology
- 73. Type of culture media: Selective Media Example: Lewenstein Jensen Medium Microbe: Mycobacterium tuberculosis
- 74. Muller Hinton Agar Used for antibiotic susceptibility testing
- 75. Dr. Nabil El Aila General Microbiology Capsule stain
- 76. Dr. Nabil El Aila General Microbiology Negative stain
- 77. Dr. Nabil El Aila General Microbiology Spore stain
- 78. Dr. Nabil El Aila General Microbiology Flagellar stain