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Flashcards by Princess Banana Hammock, updated more than 1 year ago
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| Question | Answer |
| Why perform a streak plate isolation? | Streak plate isolation produces isolated microbial colonies by diluting the original inoculum (bacterial sample) used on the agar plate. It aims to separate out single colony-forming units (CFUs) in order to allow for the growth of individual, separate colonies during incubation. It may be used in order to better visualise and quantify the morphology (size, shape, colour, and other visible attributes) of an individual colony of each bacterium, in order to help to identify the bacterium in question. Where the original bacterial sample contains more than one type of bacterial colony, streak plate isolation is indicated in order to separate out these colonies and ensure that each different type of bacterium present can be studied individually (using a 'pure sample' of each separately). Use of a 'pure' bacterial sample is the only way to obtain valid and accurate results when following-up the streak-plate isolation method with further biochemical tests to identify the type of bacteria. |
| How do we perform the streak plate method of isolation? | |
| Explain the blood agar test. | The blood agar test can be used to determine whether a bacterium is one which is able to cause haemolysis; that is, destruction of red blood cells (erythrocytes). This is because blood agar consists of 5% sheep's blood with general nutrient agar. The effect is actually produced by exotoxins (produced by the bacteria) called haemolysins. 3 results are possible in the blood agar test. A clear area around the bacterial growth indicates beta haemolysis, the complete destruction of red blood cells. A greenish discolouration surrounding the bacterial growth indicates alpha haemolysis, which is the partial destruction of red blood cells. Finally, if there is no change in the colour or opacity of the medium, this indicates that the bacteria being examined do not cause haemolysis in any form (this final, negative result is referred to as gamma-haemolysis). |
| Explain the MacConkey agar test. | MacConkey agar selects for enteric bacteria. It contains lactose and a pH indicator. Bacteria which are able to ferment lactose are called coliforms. The fermentation of lactose produces acid, lowering the pH of the colonies/medium, which due to the presence of the pH indicator, changes the colour of the colonies and sometimes the surrounding medium to pink. This is both a selective and differential test. The possible results are: no bacterial growth; bacterial growth without any colour change to pink; and bacterial growth with a colour change to pink. Most bacteria which can grow on MacConkey agar are gram-negative. |
| Explain the Eosin Methylene Blue (EMB) agar test. | The EMB agar test selects for and differentiates between different groups of enteric bacteria, similarly to the MacConkey test, but with different mechanisms. The dye inhibits the growth of gram-positive bacteria, so that any bacteria which grow on the agar is almost certainly gram-negative. It further differentiates between these gram-negative bacteria by testing whether they have the ability to ferment lactose, which is included in the EMB agar. Lactose-fermenting enteric bacteria will produce a greenish metallic sheen as the resulting acid leads directly to precipitation of the EMB dyes on the colony surface. A pink colouration is indicative of some, but not much lactose fermentation. Non-lactose-fermenting enteric bacteria may grow on the EMB agar but will lead to no colour change, with the colonies remaining colourless or taking on the original hue of their medium. |
| Explain the Brilliant Green agar test. | Brilliant Green agar is, ironically, orange in colour (see Plate A). It is used to identify species of Salmonella (except Salmonella typhi and Salmonella paratyphi). It contains the pH indicator 'phenol red' which changes colour to yellow where the pH of the medium is reduced (due to the production of acid by lactose- or sucrose-fermenting bacteria). It also contains the 'brilliant green' dye which inhibits the growth of most species of bacteria, both gram-positive and gram-negative. Possible results include (1)(See plate C) Red to pink-white colonies surrounded by brilliant red zones in the medium -- this indicates the presence of non-lactose-fermenting bacteria such as Salmonella spp. (2)(See plate B) Yellow to yellow-green colonies surrounded by yellow-green zones in the medium -- this indicates the presence of lactose-fermenting bacteria such as E. coli. |
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