Bacteriology Flashcards
Lecture 3
What is culture used for in diagnostic bacteriology? What is the major disadvantage of culture?
Used to find out which antibiotic to use (“SUSCEPTIBILITY TESTING”. The problem is that it takes about 24 hours to grow a bacteria + another 24 hours to conduct the tests.
How do you conduct a antibiotic susceptibility test?
Usually this is done by phenotypic methods - you impregnate agar with a microorganism and put antibiotic discs on it
With respect to culture distinguish between sterile sites and non-sterile sites?
Sterile sites e.g. blood/CSF. These are sites under which normal conditions there are no bacteria.
Non-sterile sites e.g. skin. These are sites which under normal conditions may be colonised by multiple different bacteria. This can make culture difficult, because it can be difficult to know which bacteria is causing pathology. Isolating the bacteria potentially is also a difficulty because it requires an understanding of its unique molecular biology.
How would you culture bacteria from non-sterile sites?
You can use selective agar plates. E.g. XLD for Salmonella
How would you culture bacteria from sterile sites?
You can use a non-selective agar because there shouldn’t be bacteria there anyway.
Does being confirmed positive for carrying a pathogenic bacteria mean you will necessarily have pathology?
No. People can carry pathogenic bacteria e.g. Staphylococcus but still be fine. Only want to take samples if suspecting an infection, because do not want to be misled. Interpret results based on patient in front of you
What is the challenge of using molecular techniques like PCR for identifying bacteria/their resistance patterns?
It might be easy to identify resistance genes for bacteria like MRSA where the resistance mechanism is understood and encoded by MecA gene. However, new resistance genes are always developing so the need for primers will preclude continued identification. Also the myriad of resistance genes makes this difficult for routine use.
In terms of aerobic and anaerobic bacteria what is the difference in culturing blood from adults and children?
In adults you would have different mediums for aerobes versus anaerobes. But in children the load you get per unit blood is much smaller which means you use 1 medium for both. This set-up disadvantages the anaerobes.
How does a blood culture test positive?
Have some indicator at the bottom of the tub, waste products from the bacteria will be detected and change the colour of the indicator. Once a blood cultures is confirmed positive do a Gram Stain to see if Gram positive or negative (this is relevant because it affects what antibiotics you use)
Generally speaking describe the distribution of Gram positive/negative bacteria in terms of where they infect.
GP = skin + soft tissue GN = abdomen and urinary tract
What is a chocolate agar? What is an example of a bacteria that finds it advantageous to grow on chocolate agar?
This is”cookedblood” - certain bacteria will not be able to lyse the blood cells so by cooking it you release some of the nutrients in the blood agar and let certain bacteria grow. Haemophilus influenzaegrows on chocolate agar
What is a macconkey agar?
An agar designed to grow gram negative organisms
What is the difference between Gram negative bacteria and Gram positive bacteria?
Gram positive has a thicker peptidoglycan cell wall which holds the Gram stain, staining purple
Gram negative has an outer membrane outside the cell wall which stops them from taking up the stain, if add counter-stain they will stain pink
GP is the most common
For GN + GP bacteria what 2 sugars help make up their peptidoglycan cell wall?
N-acetylglucosamine (NAG) and N-acetylmuramic acid (NAM)
present in both
What is clinical significance of the difference in cell walls between GP and GN bacteria? Give a relevant example?
Lots of antibiotics target the cell wall
GN bacteria have an outer membrane
This will protect the cell wall
E.g. vanomycin is very effective on Gram positives but can’t get past the outer membrane of Gram negatives
How do Staphylococci look under a microscope?
Divide in two and then the daughter cells divide again to form a clump of four
How do Streptococci look under a microscope?
Form chains
How do Gram negative bacilli?
Pinkish appearance due to counterstain
What is coagulase?
Coagulase is a virulence factor which helps Staphyloccocus cause infection
What does the coagulase test tell us?
Coagulase is a virulence factor important in pathogenesis
Skin commensals of low pathogenic potential are coagulase negative - usually can ignore but can be a danger e.g. if infect prosthetic material/pacemakers
In most cases coagulase negative Staphylococci found in blood cultures are contaminants (during extraction that is)
How can we classify streptococci?
Classify based on blood agar
Alpha haemolytic bacteria turn blood agar green (only partial haemolysis)
Beta haemolytic bacteria turn blood agar completely clear (complete haemolysis
How can beta haemolytic streptococci be classified?
Group A: Strep pyogenes cause skin and soft tissue infections
Group B: Strep algalactiae cause sepsis in neonates + infect diabetics (unclear why)
Does greater virulence mean greater resistance?
No. Strep pyogenes can cause nasty infections (scarlet fever/rheumatic fever) but they are not resistant to penicillin
What can an XLD agar be used for?
Detect Salmonella
Metabolism of xylose by Salmonella - subsequently decarboxylate lysine decreasing pH of agar which turns the phenol red yellow (other enteric bacteria do this)
+ produce hydrogen sulphide which produces black spots (Salmonella only)
How can you isolate Campylobacter?
It can survive at 42o - other bacteria can’t
What is TCBS used for?
Vibrio cholera isolation - turns it green
How can we calculate positive predictive value?What is the significance of a higher PPV?
PPV = number of true positives / total number of positives
A higher PPV means we can trust a positive value i.e. the more likely the positive is a true positive
What is the relationship between minimum inhibitory concentration and the breakpoint?
MIC is the minimum concentration of an antibiotic needed to kill the bacteria
The breakpoint is a cut-off for the concentration of an antibiotic needed to kill a bacteria.
If the MIC is above the breakpoint the bacteria is resistant.
What is the standard way of testing antibiotic sensitivity?
Disc transfusions - zone diameters is used to define whether the bacteria is resistant or sensitive. Will have a table that defines the zone diameter at which the breakpoint is defined
Why use phenotypic tests to test for antibiotic sensitivity why not just use sequencing?
A number of reasons, complicated interpretation, remember also that simply having a gene doesn’t necessarily mean you express it. Hence still use phenotypic tests. Culture is more sensitive e.g. in TB cf. some molecular techniques (need smaller number of bacteria)
What bacteria are routinely looked for as a cause of diarrhoea?
Salmonella, Shigella and Campylobacter key pnes
Also: C. difficile, cholera and E. coli can play a role
