Trond Erik Vee Aune wrote:
>>> Scott J. Coutts wrote:
>>> I must admit that the first thing I thought of was contamination. I
>> would incubate an uninoculates plate from the same batch too.
>>> When I streak the bacteria I only streak it at about one fifth of the
> total area of the plate. The colonies are showing up in the vicinity of
> where I have streaked, up to 1.5 cm away. On the other half of the
> plate, away from where I have streaked, no colonies are showing up. So
> if it is a contaminant (which we rarely see at our laboratory) it is
> dependent on something my new bacteria is producing and will only grow
> close to it. Otherwise I would not see this polar effect.
>>> I assume your loop or wire (if that's what you're using) is not too
>> hot when you streak out your bacteria? Otherwise, the media or
>> conditions in which you're growing them may not be suitable. Maybe you
>> could try some other combinations.
>> I use a loop of wire yes. When I streak other bacteria I never see the
> same strange behaviour, so I don't think it's too hot. Since this is a
> new bacteria I'm not sure of it's preferred media. I grow it on LB +
> glucose, it grows fine at 30 and 37 degrees, and after one night
> incubation I have nice sized colonies. It really seems like the in some
> way inhibit each others growth, so that they need a few mm of space
> between each colony. I know they inhibit growth of other bacteria, they
> are producing some sort of antibiotics.
Where did it come from? You might be able to find a better medium for it
depending on where you found it. You could try a more enriched media,
maybe HBA or something like that. Maybe also try growing them
anaerobically. It's just that often, if they dont grow very well on a
medium, then instead of getting confluent growth at the point of the
primary innoculation, you'll just get light growth or a few colonies.
>> Do you dry your plates before using them? Lots of motile bugs will
>> swim away from where you streak them (i.e. lots of spirochaetes,
>> Proteus etc) but usually they dont swim away to form discrete
>> colonies. But it may be possible. You're plate doesnt even have to be
>> actually wet - even if it is moist enough they can do it.
>> I dry the plates at 37 before streaking, so they should be dry. I
> thought of this solution, but it just seems too weird that they should
> swim for over a cm before proliferating and forming colonies. But at the
> moment I have no other explanation... Either long distance swimming or
> some way of shooting out spores (even though I couldn't see any spores
> through the microscope).
>
Remember that often bacterial spores will not look like fungal spores do
- they will be refractile or differentially staining bodies inside the
bacterial cell. They may or may not distend the cell. Have done any
staining of these cells?
I dont know of any bacteria that can 'shoot' their spores. The swimming
certainly sounds plausible, but not as discrete colonies. Proteus will
often swim a few cm away, then proliferate, then swim again. But usually
this shows as concentric rings of growth away from the innoculation point.
What do the colonies look like, and how big are they? Would it be
possible for you to post some pictures on the web somewhere, or in
alt.test (I dont think people appreciate binaries on this newsgroup).
Otherwise, I'd be happy to take a look at them if you want to email them
to me. It sounds interesting!
>> How much bigger than E. coli are they? Are they bigger, but still
>> bacteria sized (rather than yeast or fungus sized).
>> Difficult to say, I'm not very experienced with the microscope. Some
> people at my lab said they looked like Azotobacter vinelandii. At least
> double the size of E.coli, I would think. They were unicellular (I could
> see someone in the progress of dividing by binary fission) and showed no
> signs of differentiation, they all looked the same - slightly bent rods
> with polar flagella.
>
Do you know they have polar flagella just because of the motion of their
movement, or can you see them? If you can see them, then I dont think
they're bacteria. But the cell size sounds like bacteria.
>> Thank for the input Scott!
>
Anytime.
Ok, everything you replied to rules out what i said in my first post...
hm... well, I dont know of any bacteria that exhibit this behaviour. I
doubt that the cells are inhibiting their own growth. I cant see any
reason for them to have evolved such a trait (although many eukaryotic
cells will have contact inhibition that stops them from growing when in
close proximity). Certainly many bacteria can inhibit or kill other
species, but never usually their own.
I'd definitely recommend 16S sequencing if you have it available. It
will give you a quick and definitive answer (unless it's completely
unknown). Do you know about the RDP (ribosomal database project)? It is
a database of all the known (read: submitted) species ribosomal
sequences. You can view them as a (huge) tree and add your sequence to
it to see visially where it fits. If yours is unique, it will simply
join all the others (if you choose to submit it)- if you look at the
full tree there's plenty of entries that have names like "unidentified
organism from swamp water" hehehe.
Anyhow, here's the link if you're intersted:
http://rdp.cme.msu.edu/html/
PS: Do you mind me asking how you came across this bug? I've sequenced a
few contaminants from plates we've found in our storage in the fridge,
just for the fun of it. Sometimes it's interesting (:
--
Scott J. Coutts
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Bacterial Pathogenesis Research Group
Department of Microbiology Ph [+61 3 9905 4838]
PO Box 53 Fax [+61 3 9905 4811]
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