Interpreting gram stain results...unknown organism

[fansounds]

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

 

[Ygggdrasil]

Is it possible that the staining is incomplete (i.e. not enough of the crystal violet was present to stain the gram positive bacteria completely)? Wikipedia suggests that some strains of bacteria can give variable results in gram staining: https://en.wikipedia.org/wiki/Gram_staining#Gram_variable_and_gram-indeterminate_bacteria

 

[fansounds]

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

 

[BillTre]

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.

 

[fansounds]

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

 

[BillTre]

If everyone else has the same problem, it could be the stain.

If not I would guess the decoloring.