1. IDENTIFIKASI BAKTERI OLEHSUDRAJAT FMIPA UNMUL 2009

2. A. KARAKTERISTIK FENOTIF Metabolic differences Colony morphology –Colonies can exhibit macroscopic differences e.g., Colonies of streptococci generally form fairly small colonies e.g., Colonies of Serratia marcescens produce a pigment and are often red when incubated at 22 o C e.g., Colonies of Pseudomonas aeruginosa often produce a soluble greenish pigment

3. A. KARAKTERISTIK FENOTIF Metabolic differences Culture characteristics –Selective and differential media can aid in the identification of microbes Selective media favors the growth of certain types of microbes by inhibiting the growth of others Differential media contains a substance that certain bacteria change in a recognizable way

4. A. KARAKTERISTIK FENOTIF Metabolic differences Culture characteristics –MacConkey agar is both selective and differential Bile salts and dyes inhibit all but certain gram- negative rods –“Selective” Acid produced by bacteria able to ferment lactose will turn a pH indicator red and form red colonies –“Differential”

5. A. KARAKTERISTIK FENOTIF Metabolic differences Culture characteristics –Blood agar can be used to detect bacteria producing hemolysins e.g., Harmless Streptococcus species residing in the throat often cause alpha-hemolysis –Greenish clearing around colonies e.g., “Strep throat”-causing Streptococcus pyogenes causes beta-hemolysis –Clear zone around colonies

6. A. KARAKTERISTIK FENOTIF Metabolic differences Culture characteristics –Media lacking nitrogen can be used to detect nitrogen-fixing bacteria e.g., Azotobacter can be identified from soil samples incubated aerobically on such media

7. A. KARAKTERISTIK FENOTIF Metabolic differences Biochemical tests –Generally necessary for more conclusive identification –Most rely on pH indicator or color change when a compound is degraded

8. A. KARAKTERISTIK FENOTIF Metabolic differences Biochemical tests –Sugar fermentation e.g., Lactose, sucrose, glucose, etc. Fermentation results in acid production –pH indicator changes color –Pink  yellow Inverted tube (Durham tube) collects any gas produced

9. A. KARAKTERISTIK FENOTIF Metabolic differences Biochemical tests –Urease detection Enzyme degrading urea –Urea  CO 2 & NH 3 pH indicator turns bright pink in alkaline conditions Helicobacter pylori can be detected using a breath test

10. A. KARAKTERISTIK FENOTIF Metabolic differences Biochemical tests –Urease detection Helicobacter pylori can be detected using a breath test –Causative agent of most stomach ulcers –Culturing not necessary –Patient drinks solution containing 14 C-labeled urea – 14 C in expired are indicates presence of urease

11. A. KARAKTERISTIK FENOTIF Metabolic differences Biochemical tests –Catalase Commonly occurring enzyme –Possessed by most bacteria growing in the presence of oxygen –Absent in lactic acid bacteria »e.g., Streptococcus »Beta-hemolytic catalase-negative bacteria from a throat culture may be Streptococcus pyogenes

12. A. KARAKTERISTIK FENOTIF Metabolic differences Biochemical tests –Catalase Simple assay –H 2 O 2  H 2 O & O 2 –O 2 bubbles are visible

13. A. KARAKTERISTIK FENOTIF

14. Organisms are identified using a dichotomous key –Multiple biochemical and other tests are typically required Multiple tests are generally run concurrently –Avoids waiting for incubation time for each test

15. A. KARAKTERISTIK FENOTIF Metabolic differences Biochemical tests –Commercial modifications of traditional biochemical tests e.g., API TM system, Enterotube TM

16. Metabolic Phenotypic Exam cultural approaches –required for positive diagnosis of infection –isolation and ID of pathogen –accuracy, reliability, and speed –specimen collection is important –commonly used for positive identification of most prokaryotes methods used include –culture characteristics –biochemical reactions process

17. Metabolic Phenotypic Exam culture characteristics –organisms grown in a pure culture are easier to identify due to the high number of organisms obtained E. coli

18. Metabolic Phenotypic Exam culture characteristics –use of selective or differential media can provide additional information selective media inhibits the growth of organisms other than the one being sought differential media contains substances that particular bacteria change in a recognizable way

19. Metabolic Phenotypic Exam When identifying a suspected organism, a series of differential media is inoculated. After incubation, each medium is observed to see if specific end products of metabolism are present. This can be done by adding indicators to the medium that react specifically with the end product being tested, giving some form of visible reaction such as a color change. The results of these tests on the suspected microorganism are then compared to known results for that organism to confirm its identification.

20. Metabolic Phenotypic Exam Some bacteria will produce the enzyme catalase. Catalase will break down hydrogen peroxide releasing oxygen, which is indicated by the bubbles that have formed.