orgs and their characteristics

some info for your little book::
H2S producers
Salmonella
Proteus

non motile orgs
Klebsiella
Shigella

lactose fermenters
Escherichia coli
Enterobacter aerogenes
Klebsiella pneumoniae, oxytoca


non lactose fermenters
Salmonella enteritidis, typhimurium, typhi, species
Proteus vulgaris, mirabilis
NB! I have used incorrect nomenclature rules in naming the above. Salmonella has many species, which I have separated by commas, same with Proteus.

Please remember to underline and correct spelling.

CLED

CYSTEINE LACTOSE ELECTROLYTE DEFICIENT AGAR
This agar is used to differentiate between urinary orgs. The deficiency in lectrolytes prevents the swarming of Proteus species, a GNB and a member of the coliforms. The medium contains lactose which serves as a CHO (carbohydrate) source.Therefore we can get lactose fermenters - yellow colonies- and non lactose fermenters - blue colonies.

You must know what CLED stands for, the purpose of the medium and the colours of the 2 types of orgs.

survival of the fittest

also known as: how to survive BMT3!!!

Keep your ears and eyes open, for news about changing timetables, pracs and lecture times. Always keep a notebook handy for scribbling down important info you may not be aware of (and your friend does!). Get and keep on the best side of all lecturers. Ho you ask??? Firstly show respect by greeting, smiling, and paying attention when they are talking. Listen and appear as if you are. Ask questions. If you have a schedule, read up on the forthcoming lecture so you can clear any doubts during the lecture itself. Don't be a smart-ass, it does not help in the long run. Speak up for yourself, no one else will. Always complete any homework given to you. Show that you care, and the lecturer will take care of you. Show that you don't care, and he will end up ignoring you.
Do some reading on your own. All the subjects are new, and require getting to grips with. Get your tongue around the new terms. Explain things to yourself. and to others. Use the lab time available to its maximum. Yes you are hungry and tired, but next week is another prac, and today will be history. You will not get another chance to look at all aspects of the techniques or to ask questions of the lecturer.

enough already. over and out

nitrate reduction test

the nitrate broths are inoculated with the organism and incubated aerobically, 18-24 hours at 37 degrees.

after removing from the incubator, add solution A and B (in that order, 5 drops each) to each tube. Wait for a few minutes for the colour to develop. A red colour indicates a positive reaction, i.e. reduction of nitrate to nitrite.

zinc is added to all negative tubes. Zinc is able to reduce nitrates to nitrites, therefore a development of a red colour here indicates a negative reaction.

the tube that still shows no colour change is..... wait for it........ positive for nitrate reduction.

why you ask??????????

simply silly

because the organism reduced nitrates to ammonia gas. You could try smelling it to be sure, but I wouldn't advise it. Apart from fainting and becoming a hazard on the floor, (it's nasty down there), I can't see myself picking you up. Duh!!!!

isn't all this stuff amaaaazing!!!!

success in micro1 practical examinations

I think you and I both need this.

So what does one need to do in order to survive the prac exam?????

When you come into the lab, stop talking. You are already stressed out so you don't need to become more stressed by annoying the staff. You should have already taken out your pen or pencil, marker and labcoat, but only put on the labcoat in the lab. Don't spend time digging in your bag. Get to your place as quickly as possible.

Read through the test paper quickly and determine if you have the correct paper and all relevant documentation. Write down your name and student no.

Listen to any instructions from either the lecturer or lab technician. Check that you have heard, understood and know where all the media, cultures and apparatus are located. Ensure that you know where all the demos are positioned and have heard the instructions for the demos.

You need to organise the time available to you appropriately. Start with the procedures that require the most amount of time.

I need to organise my thoughts on this issue. Back later.

TSI contd....

I forgot to mention yesterday that the sugars in TSI are in different quantities, for a reason. Glucose is incorporated at 0.1% while lactose and sucrose are present in 1%. This is so that the orgs are forced to utilise lactose and/or sucrose after glucose is used up.

There is a systematic way of reporting the results. Remember that we are looking at and reading 4 different results, viz, acid or alkaline production in both the butt and slant, gas production and H2S production. We use the abbreviations A for acid and K for alkaline. Acid production is shown by a change in colour from red to yellow, while a change in colour to a deeper red is indicative of alkaline reaction.

All coliforms will give a positive result for glucose utilization, therefore an acid butt.

You must report acid or alkaline production in slant or butt, gas production and H2S production. An exemplar is A/A/no gas/no H2S.

Remember to give me some kind of indication of the order in which you are reporting.

see you!

TSI

Here is one of my all time favourite media, triple sugar iron. Used to differentiate between coliforms on the basis of utilization of glucose, lactose and sucrose, gas production and H2S production. The medium is an agar slant dispensed in 10 ml amounts. It is pale red in colour before inoculation.

It contains 3 carbohydrates or sugars, glucose, lactose and sucrose. Glucose is found in the butt (bottom of slant) and lactose & sucrose at the slant. utilization of any sugar results in a change in colour from pale red to yellow, which indicates acid production. An alkaline reaction is indicated by a deeper red colour.

H2S production is seen by a blackening of the medium. Gas production can be detected by any one of 3 ways:

bubbles along the tube

cracks in the medium

the entire medium moves up in the tube.

All coliforms utilize glucose. Actually if orgs had a choice between many sugars they would use glucose first, since it is easier to digest (degrade). Once the glucose has been used up, the org moves to the slant to the lactose and sucrose. If the org is a LF it utilises the lactose and sucrose; if the org is a NLF, it does not.

You can deduce the reaction an org would give on another medium based on its reaction on TSI. Try it yourself.

next time: expected reactions for coliform on TSI

format of reporting results of TSI

Lowenstein and Jensen agar

This is a mouthful!

used to culture and differentiate between species of Mycobacterium. It is said to be egg based because it uses a lot of homogenized eggs as its base. Also added are many inhibitors of growth. Mg sulphate and Mg citrate inhibit the growth of all other orgs except Mycobacterium. Malachite green inhibits any contaminants. Cycloheximide is an antifungal whilst lincoymcin and naladixic acid are antmicrobial.

You may well wonder why so many ways to prevent growth of other orgs. Simpe really. Culture of Mycobacterium requires incubation of up to 6 weeks. Imagine if both other microbes and fungi could grow uninhibited in this time. We would have to search high and low, narrow and wide, long and short to find the Mycobacteria. Remember that both fungi and regular ol' bacteria only require 18 to 24 hours to produce adequate growth. Another reason for the inhibitory substances relates to the specimen of choice, the sputum. The specimen would contain many normal flora along its route of expectoration. We don't want them!

Now the medium is pale green in colour, taken from malachite green which inhibits contaminants. It is not an agar plate, but a agar slant dispensed in small bottles with a screw top lid.

MSA

Change to microbiology here. Hope its OK?

Today we discussed media used in medical microbiology for culturing, isolating and identifying microbes of medical importance. Here is a short review of what we learnt.

MSA also known as mannitol salt agar.

used to distinguish between Staphylococcus species on the basis of mannitol fermentation, when isolated from foods, milks and clinical specimens.

The ph indicator is phenol red.

The NaCl content is moderate at 7.5 %, which makes this medium selective because it inhibits non halophiles. A halophile is an organism that is able to grow in a medium that has an increased NaCl concentration.

so this is what happens:

the organism grows, utilises the mannitol, produces waste products which in turn change the ph of the medium. This causes the ph indicator to cause a change in the colour of the medium from the original pale pink or red to yellow. Mannitol fermenting orgs either grow as yellow colonies or produce a yellow zone around the colonies. Non mannitol fermenting orgs do not utilise the mannitol therefore producing pink or red colonies. There is no associated colour change of the agar around the colonies.

can you determine the reactions shown by Staphylococcus aureus and S epidermidis?

cell markers

Lets talk about cell markers. These are "addresses" of the cells so that other cells can identify them.

Lets imagine this, ok? Say you attend a particular college or university. How would you recognize a student who attends the same? She or he would probably be travelling the same way, or be wearing the same t shirt with the college logo on it. How would you recognize a student who attends the same collge, but is registered for a programme in another faculty? Probably from the fact that you see him or her during the breaks, but not in one of your classes. Ok, so far, so good.

What about recognizing a student studying the same programme as you, but not a peer? Perhaps by their harried worried look if they are ahead of you.

By the above explanation, you can see that we use visual clues to identify people, even though we may not know them. Likewise, in the immune system, cells do not have their names printed on their surfaces, but they are still recognized by other cells in the vicinity. This is achieved by cell markers.

Now cells express all sorts of cell markers, for many different reasons. They are used to indicate the lineage of the cell, whether the cell is a naive or activated cell, or whether they are immature or mature.