Bacteriology Flashcards
What are the common bacteriology diagnostic techniques? (x4)
□ CULTURE (agar plates): either from sterile sites such as blood and CSF, or non-sterile sites such as skin where commensal bacteria are found. This is the most common! □ SEROLOGY: study of the blood serum to identify body response to infection – antibody testing. □ MOLECULAR TECHNIQUES: such as PCR, electrophoresis, arrays, blotting. □ ANTIMICROBIAL SUSCEPTIBILITY TESTING: phenotypic (meaning can be studied physically in front of you; does not involve studying genes) method of testing susceptibility of bacteria to certain treatments e.g. antibiotic sensitivity using antibiotic discs.
Why do we continue to investigate bacteria in culture?
Seems like a primitive method, but growing bacteria in culture is really good for investigating the effect of drugs/stimuli on the bacteria – ANTIMICROBIAL SUSCEPTIBILITY TESTING. In many cases, we don’t just want to know what bacterial infection a patient has got, but also how to treat it. Culturing bacteria means that we can investigate whether to use an antibiotic or approach treatment differently because of resistance.
Why are molecular techniques not often used in bacteriology?
Molecular methods tell us what bacteria the sample is. This is not what we are most interested in – we are more interested in how we can treat the infection: With a specific antibiotic or a general antibiotic because the infection has resistance? In molecular techniques, in order to know whether a bacterium is antibiotic resistant, you would need to be able to identify SNPs in the genome that give the bacteria the resistance. However, there are 1000s of genes involved in resistance, so this method is too inefficient/unreliable right now. Phenotypic methods such as culturing the bacteria and applying antibiotics to test sensitivity is most efficient.
What is the definition of sterile?
Free from bacteria or other living microorganisms.
What is the first step for blood samples sent to a bacteriology lab?
□ Blood cultures placed in bottles with growth medium. □ One bottle promotes aerobic respiration, whereas the other forces anaerobic respiration. □ Broth inside contains nutrients for bacteria and sample is intubated at 37 degrees. □ Broth and sample are agitated to ensure broth circulates round all the sample. □ Bottom of the bottle contains an indicator. As the bacteria grow, they create toxic metabolites which interact with the indicator which changes colour. □ Machine will flag colour changes as an indicator that bacteria are present. Remember, blood is a sterile medium so shouldn’t have bacteria. □ Usually takes between 16-20 hours.
What happens to blood cultures that are tested positive for bacteria?
□ Bacteria is identified and a diagnosis is made. The first step in identification is GRAM STAINING – determines whether the bacteria is Gram positive or Gram negative. □ Shape and arrangement of bacteria is studied under a microscope. □ More specific tests are carried out after these initial tests to identify the specific bacteria in the blood sample. □ Blood cultures are sent for antimicrobial sensitivity testing on agar plates.
What are the key differences between Gram positive and Gram negative bacteria?
Their cell walls differ! Gram positive bacteria have a thick peptidoglycan cell wall and a single cytoplasmic membrane. Gram negative bacteria have a thin peptidoglycan layer and a lipopolysaccharide outer layer as their cell wall.
How do Gram positive and Gram negative bacteria stain?
□ Gram positive stains deep purple – complex of crystal violet and iodine get stuck in the thick peptidoglycan wall. □ Gram negative do not stain with the primary stain. They are stained pink/red by a counterstain, commonly safranin.
Why is it important to differentiate the Gram staining of bacteria?
Antibiotic treatment differs between the two.
What are the two main groups of Gram positive bacteria? How can these be identified based on shape and arrangement?
□ STAPHYLOCOCCI are spherical (cocci) and divide, forming bunches like grapes.
□ STREPTOCOCCI are spherical (cocci) and divide end on end to form chains.
How are staphylococci bacteria further investigated in a bacteriology lab? What is the significance of coagulase?
□ Staphylococci are split into two groups using a coagulase test which tests the ability of the bacteria to form a clot in plasma. □ They are coagulase positive if they can clot, and coagulase negative if they cannot. □ Coagulase tells us about the pathogenicity/virulence of the staphylococci. Presence of coagulase increases the power of the bacteria to infect.
What are coagulase positive and negative staphylococci?
□ Staphylococcus aureus (including MRSA) is coagulase positive. □ Coagulase Negative Staphylococci are skin commensals of low pathogenic potential. Their presence is usually caused by mixing of skin commensals with blood in venepuncture.
When can coagulase negative staphylococci become problematic?
Can infect prosthetic material (e.g. hip replacements) causing line, pacemaker infections and endocarditis.
How are streptococci further investigated in a bacteriology lab?
□ Bacteria are cultured on blood agar plates – agar plates which contain blood as the medium (usually from a sheep or horse).
□ Streptococci can be distinguished depending on whether they perform alpha, beta or gamma haemolysis.
□ Alpha: incomplete haemolysis i.e. turns agar green. Includes Strep. Pneumoniae (cause of pneumonia and meningitis).
□ Beta: complete haemolysis i.e. clears the agar. Includes Strep. Pyogenes (skin and soft tissue infection) and Strep. Agalactiae (neonatal sepsis).
What, in clinical practice, can pose a challenge to culturing bacteria in a bacteriology lab?
If a clinician suspects a patient has a bacterial infection and administers antibiotics before a blood sample is taken, once blood is taken and sent to bacteriology, the laboratory could underestimate the infection present in the patient when it is tested.
