ANSC20003 Chap.12 From Milk Sample to Named Pathogen
From Milk Sample to Named Pathogen
One quarter, one sample, two weeks
Mastitis in the dairy cow is inflammation in one quarter of the udder or in several, and most cases are intramammary infections with pathogens entering through the teat canal. It is called clinical or subclinical according to whether anything can be seen, and where signs are absent a high somatic cell count indicates an immune response.
Determining the causative agent is central to managing herd health, and these two sessions do exactly that.
Each student receives an individually numbered milk sample from a single quarter of a cow suspected of mastitis, and the first assignment is written from that sample's results.
Four plates, four jobs
The milk sample is the primary inoculum for streak dilutions on three plates: a medium selecting salt tolerant organisms and differentiating on mannitol fermentation, a medium selecting enteric organisms and differentiating on lactose fermentation, and an enriched medium differentiating on haemolysis and aesculin hydrolysis.
A fourth plate is used for a handwashing exercise with clean and dirty halves, incubated alongside the others and compared the following week. That exercise is not an aside: normal flora from one animal can be a source of infection in another, and hand hygiene is part of preventing mastitis spreading through a herd.
Describe before you interpret
Colony description is a protocol.
Measure the diameter, describe pigmentation, then form, elevation and margin, then surface as smooth, rough or mucoid, then opacity and texture. The stained slide gets the same treatment: whether one or several organism types are present and which predominates, the staining characteristics, the shape, the size and the configuration.
Each plate then needs a second sentence saying what growth on that medium means, and omitting the interpretation is the most frequent gap.
From a single colony to a presumptive identity
The Gram smear is prepared from one typical, well isolated colony, dried, heat fixed and stained through a fixed sequence in which the decolourisation step is short and unforgiving.
The catalase test detects an enzyme by adding hydrogen peroxide to a colony lifted without agar, since blood-containing media give a false positive. A diagnostic key then arranges the plate results, the Gram reaction, the cell arrangement and the biochemical result into a presumptive identification.
What this chapter covers
- 01
Mastitis as an intramammary infection, and why the causative agent is worth naming
- 02
Streak dilutions on selective and differential media from one quarter sample
- 03
The handwashing plate as an experiment with its own control
- 04
The colony description protocol and its permitted terms
- 05
The stained cell descriptors, including the configuration people omit
- 06
The Gram stain sequence, and the step that reverses a result
- 07
The catalase test, its false positive, and how a diagnostic key is read
Resolving a result set that contradicts itself
- 1Explain why the three results do not sit together.
- 1Name the most likely technical cause.
- 1State the check, and the order in which conclusions should follow.
Key terms
- Streak dilution
- A plating technique that separates cells across successive regions of a plate so that single colonies can be obtained.
- Colony morphology
- The described appearance of a colony across diameter, pigmentation, form, elevation, margin, surface, opacity and texture.
- Haemolysis
- The pattern of red cell breakdown around a colony on blood-containing medium, read as one of several described types.
- Heat fixing
- Passing a dried smear briefly through a flame so that cells adhere to the slide and survive staining and rinsing.
- Catalase test
- A biochemical test in which hydrogen peroxide is added to a colony, with bubbling indicating that the organism produces catalase.
From Milk Sample to Named Pathogen FAQ
Why must the Gram smear come from a single colony?
Because every descriptor after the smear assumes one organism. A smear taken across mixed growth puts more than one organism on the slide, so predominant type, shape, size and configuration all describe a mixture and none of them can be attributed.
Why is agar avoided when picking a colony for the catalase test?
Red blood cells contain catalase, so carrying blood-containing agar onto the slide produces bubbling that comes from the medium rather than from the organism. The result would be a false positive that then propagates into the identification.
What is the handwashing plate actually testing?
It compares an unwashed hand with a washed and dried hand on two halves of one plate, so the comparison is internal. The following week you count colony morphologies on each half, look for any resembling those from your milk sample, and judge whether number or type reduced.
How far does the diagnostic key take an identification?
To a presumptive identification. Every input it uses, plate results, Gram reaction, cell arrangement and a biochemical test, is phenotypic, so confirmation to species would require further biochemical testing or sequencing.
Exam move
Rehearse both description vocabularies until you can produce them without prompting, since the practical report and the assignment are marked on completeness rather than on inference. Learn the Gram sequence with its timings and treat decolourisation as the step to watch.
Practise writing a two-sentence entry for each plate, one describing and one interpreting, because that pairing is what converts observations into an identification and it is the gap most often left in submitted reports.
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