Interpreting gram stain results...unknown organism

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Discussion Overview

The discussion revolves around interpreting gram stain results for an unknown organism in a microbiology homework assignment. Participants explore the implications of observing both purple and pink cocci in the staining process, questioning the reliability of the results and the potential for staining errors.

Discussion Character

  • Homework-related
  • Debate/contested
  • Exploratory

Main Points Raised

  • One participant notes confusion over observing both purple and pink cocci, questioning whether to rely on the majority color to determine gram positivity or negativity.
  • Another participant suggests that incomplete staining could be a reason for the mixed results, referencing variability in gram staining outcomes for certain bacterial strains.
  • A participant asserts that their professor indicated the results are not due to gram variable or gram indeterminate bacteria, leading them to consider redoing the stain.
  • Concerns are raised about the quality of the staining process, with suggestions that old or poorly made stains could lead to erroneous results.
  • Participants discuss the importance of proper washout steps and techniques, such as centrifuging or filtering, to improve staining accuracy.
  • One participant expresses that their issues may stem from poor technique, specifically regarding the decolorization process, and plans to redo the experiment.

Areas of Agreement / Disagreement

Participants express uncertainty about the reliability of their gram stain results, with some agreeing that poor technique or staining quality may be contributing factors. However, there is no consensus on the underlying cause of the mixed results, and multiple competing views remain regarding the interpretation of the findings.

Contextual Notes

Limitations include potential dependencies on staining techniques and the quality of reagents used, as well as unresolved questions about the specific characteristics of the unknown organism.

fansounds
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Homework Statement


We've been given unknowns and now have to determine what the organism is. I'm having trouble interpreting gram staining results. As far morphology they are all cocci and (sort of) in clusters but (this is the confusing part) they are both purple and pink. I understand that purple indicates gram+ and pink is gram- , but there should be one genus of organism not two. So shouldn't there be only purple or only pink, but not both? Should I be looking for the "majority" color and using that to decide whether I have pos or neg? That is, if its mostly purple, does that mean pos, ...but if so, then why are there still pink cocci on the slide?

Homework Equations


I also understand why gram-pos appear purple, --that the crystal violet-iodine complex is retained due to thicker cell walls in gram-pos... and thinner cell walls in gram-neg, so they are decolorized and appear the color of the counterstain.

Also I definitely did not see any rods in my slide, just cocci.

The Attempt at a Solution

 
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thanks for answer ygggdrasil...its definitely not gram variable/gram indeterminate because I asked prof about this...I think I will redo the stain though
 
Are others having similar problems?

Old or poorly made stains can give erroneous results.
Sometimes it is useful to centrifuge or filter them (using something like a micron scale filter) to remove particles that can adhere to tissue non-specifically.

Washout steps can be important for getting a good differentiation in staining.
 
yup centrifuging would help but the thing is this is just an intro to microbio class so I'm restricted to what test I can do. So this is all probably due to poor technique (specifically either over/under decolorizing) so I'm just going to redo
 
If everyone else has the same problem, it could be the stain.

If not I would guess the decoloring.
 

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