Chapter 43

1. Chapter 43 Basic Microbiology

2. The Medical Assistant’s Role in the Microbiology Laboratory • Preparing cultures • Allow bacteria to grow at least 12 hours before examining culture • Sensitivity identifies which antibiotics will kill microorganism causing infection

3. The Medical Assistant’s Role in the Microbiology Laboratory • Use exact technique to avoid laboratory error • Use sterile equipment • Send culture to laboratory in reasonable amount of time

4. The Medical Assistant’s Role in the Microbiology Laboratory • Identification of organisms done successfully within 24–72 hours

5. Microbiology • Bacteria are found naturally in the body • Normal flora • Always present and help with immune system • Pathogens cause disease

6. Microbiology • Classification • Taxonomy deals with classification of living organisms • Carolus von Linnaeus devised current classification system • No universal agreement on one system

7. Microbiology • Classification • Kingdoms • Plants • Animals • Protists • Prokaryotes (lower protists) • Eukaryotes (higher protists)

8. Microbiology • Nomenclature • System for naming bacteria • Genus • First name; capitalized • Species • Second name; not capitalized

9. Microbiology • Nomenclature • Bacteriologists and microbiologists • Parasitology • Virology • Mycology • Reference laboratory • Report certain types of bacteria and yeasts to Department of Public Health

10. Microbiology • Cell structure • Basic bacterial cell >> • DNA (deoxyribonucleic acid)

11. Equipment • Autoclave • Used to sterilize equipment • Not used with presterilized and disposable equipment

12. Equipment • Microscope • Used to view organisms that cannot be seen with naked eye • Prepared slides used

13. Equipment • Safety hood • Aerosols can be released into air when culturing and are potentially dangerous if inhaled • Use of hood is mandatory when performing culture on specimen with potential aerosol • Used to minimize odors

14. Equipment • Incubator • Has constant temperature of 35–37°C • Grows aerobic or anaerobic organisms • When culturing, set up some cultures in oxygenated environment as well as oxygen-reduced environment

15. Equipment • Anaerobic equipment • Absence of oxygen to grow anaerobic bacteria • Use of candle jar • Gas pack jar • Specimens sent to reference laboratories • Gram stain used to observe gross morphological features of bacteria

16. Equipment • Inoculating equipment • Inoculating loop • Inoculating needle • Stab culture used for certain biochemical tests used for identification

17. Equipment • Incinerator • Quickest method of sterilization • Electrical incinerator or Bunsen burner • Media • Host of substances • Used to foster growth of bacteria

18. Equipment • Refrigerator • Used to store materials • Temperature of 2–8°C • Never store food or drink or medication with specimens

19. Safety When HandlingMicrobiology Specimens • Personal protector when handling microbiology specimens • Wear PPE at all times • Remove when leaving for the day • Buttoned laboratory coat or apron, safety goggles, and gloves

20. Safety When HandlingMicrobiology Specimens • Personal protector when handling microbiology specimens • Use of hood or shield • Never eat, drink, smoke, or put objects into mouth • Do not touch contact lenses or apply makeup • Wash hands frequently

21. Safety When HandlingMicrobiology Specimens • Work area • Use strong germicide before and after daily use or immediately after spills • Dust-free and clean at all times • Uncluttered • Avoid body burns or files

22. Safety When HandlingMicrobiology Specimens • Specimen handling • Check for leaks and contamination on containers • Wear gloves • Use appropriate container • Handle all specimens as if contaminated

23. Safety When HandlingMicrobiology Specimens • Disposal of waste and spills • Biohazard symbol • Separation of biohazardous wastes • Disinfect spills with 10 percent bleach solution

24. Quality Control • All equipment with temperature controls should be monitored daily • Microscopes should be cleaned and kept dust-free • Media of all types should not be used past shelf life • Should be stored at proper temperatures • Checked for growth with known organisms for quality control

25. Quality Control • Procedure manual with all standard operating procedures written down should be updated periodically • Many microbiology laboratories subscribe to associations that periodically send unknown samples to be set up and identified

26. Collection Procedures • Check to see if culture was: • Collected properly • Delivered within a reasonable period of time • Collected in sufficient quantity

27. Collection Procedures • Common microbiology specimen sites • Place in appropriate container • Bring to laboratory • Rejecting specimens

28. Collection Procedures • Factors determining successful isolation of causative pathogens • Proper collection from infection site • Collection of specimen during infection period • Sufficient amount of specimen • Appropriate specimen container • Appropriate transport medium

29. Collection Procedures • Factors determining successful isolation of causative pathogens • Specimen labeled properly • Specimen brought to the laboratory in a minimal amount of time • Specimen collected before administration of antibiotics • Specimen inoculated onto proper media and placed in correct atmosphere to ensure growth

30. Specific Collection Requirements • Urine • Collecting a clean-catch specimen • Use of catheterization • Nose • Nasal-pharyngeal swab collects specimen • Place swab in sterile tube for transport to laboratory

31. Specific Collection Requirements • Throat • Use sterile tongue depressor to hold patient’s tongue down • Avoid swabbing sides of mouth and tongue

32. Specific Collection Requirements • Wound • Use of sterile needle or swab to aspirate pus-filled fluid from wound • Use of anaerobic transport medium

33. Specific Collection Requirements • Sputum • Patient coughs deeply and expectorates into sterile container • Should be morning specimen • Use of special container

34. Specific Collection Requirements Click Here to play the video

35. Specific Collection Requirements • Stool • Ova and parasites • Bacterial cultures • Non-sterile containers • Contamination of urine

36. Specific Collection Requirements • Cerebrospinal fluid (CSF) • Lumbar puncture • Fluid dispersed in several departments of clinical laboratory • Use of incubator • Refrigeration can kill meningitis-causing bacteria

37. Specific Collection Requirements • Blood • Development of septicemia • Collection of cultures • Variety of collection devices available

38. Bacterial Shapes Cocci Bacilli Spirilla

39. Microscopic Examination of Bacteria • Dyes • Derived from coal tar • Acidic dyes carry a negative ion • Basic dyes carry a positive ion • Methylene blue binds to DNA and RNA of cell

40. Microscopic Examination of Bacteria • Simple stain • Uses single stain on fixed slide for given period of time • Shows structure and arrangement of bacterial cell • Takes no more than 3 minutes to stain • Gives little information other than size and morphological arrangement

41. Microscopic Examination of Bacteria • Differential stain • A common differential stain is the gram stain • Use of decolorizer and counterstain • Developed in 1884 by Dr. Hans Christian Gram • Differentiates bacteria by gram stain ability of being negative or positive • Use of gentian or crystal violet reagents

42. Microscopic Examination of Bacteria • Differential stain • Identifies gram-positive and gram-negative bacteria • Staphylococcus • Streptococcus • E. coli • Proteus • Morphological arrangement, shape, and gram-stain characteristic help identify bacteria

43. Microscopic Examination of Bacteria • Acid-fast stain • Specific stain • Allows microscopic examination of acid-fast mycobacteria • Use of heat or powerful dye • Ziehl-Neelsen stain • Kinyoun stain

44. Microscopic Examination of Bacteria • Special techniques • Used when flagella, spore, capsule, or nuclei of cells are present • Tests without staining • Wet slide preparation • Hanging drop

45. Microscopic Examination of Bacteria • Potassium hydroxide (KOH) preparation • Used for study of fungi and spores • Fragments of human hair, skin, or nails placed on slide with drop of 10 percent KOH and coverslip • KOH clears debris

46. Microscopic Examination of Bacteria • Potassium hydroxide (KOH) preparation • Set slide at room temperature for one-half hour before examination for debris settlement • Use of phase or dark-field microscope • Dispose of properly (live organisms) • Direct microscopic examination of culture and infectious bacteria

47. Culture Media • Inoculate material on proper medium for growth • Reliability of results • Fastidious bacteria need specialized medium to grow • Aerobic bacteria grow only in oxygen

48. Culture Media • Common bacteria and growth requirements • Transport media • Can be solid, liquid, or semisolid substance

49. Culture Media • Contains nutrients to support growth of bacteria • Vitamins • Sugar • Salt • Minerals • Amino acids • Addition of special products

50. Culture Media • Agar • Solid media • Poured in petri dish or tubes • Broth tubes store semisolid media